Value chain hygiene practices and microbial contamination of street and market vended ready-to-eat grasshopper, Ruspolia differens in Uganda: Implications for food safety and public health.

Food safety is a major public health issue particularly in developing countries. Ready-to-eat street-vended foods contribute significantly to dietary intake in urban and peri-urban areas, but with elevated public health risk. In this study, hygiene and food safety practices as well as the microbial contamination in Uganda's edible grasshopper value chain were evaluated."A total of 29 grasshopper-processing households participated, and grasshopper samples collected. Indicator pathogens were analyzed using standard microbiological methods. In Kampala 50% and in Masaka 12% households had earth floors. All households in Kampala were one or two-roomed dwellings with no separate room as a kitchen, and shared a toilet. In contrast, 59% of households in Masaka had three or more rooms, 35% had a separate room for a kitchen and 47% did not share a toilet. 83% households in Kampala and 56% in Masaka obtained drinking water from public taps. Handwashing was inadequate and none of the actors was observed to wash their hands after taking a break or handling waste. For vendors, wearing protective clothing was not common, with only 28.5% in Kampala and 30.8% in Masaka wearing an apron. Containers for vending grasshoppers were largely uncovered and the utensils for measuring the grasshoppers were left mainly uncovered. Indicator organisms, Escherichia coli and Salmonella typhimurium, were detected. E. coli was the most common contaminant, but with lower levels in Masaka compared to Kampala. S. typhimurium was mainly a burden in Kampala. Our findings demonstrate that there are enormous contributors to poor hygiene and sanitation along the edible grasshopper value chain. The existence of pathogenic bacteria such as E. coli in ready-to-eat foods imply that their consumption poses a health risk.

Differences in gene expression profiles in early and late stage rhodesiense HAT individuals in Malawi.

T. b. rhodesiense is the causative agent of Rhodesian human African trypanosomiasis (r-HAT) in Malawi. Clinical presentation of r-HAT in Malawi varies between foci and differs from East African HAT clinical phenotypes. The purpose of this study was to gain more insights into the transcriptomic profiles of patients with early stage 1 and late stage 2 HAT disease in Malawi. Whole blood from individuals infected with T. b. rhodesiense was used for RNA-Seq. Control samples were from healthy trypanosome negative individuals matched on sex, age range, and disease foci. Illumina sequence FASTQ reads were aligned to the GRCh38 release 84 human genome sequence using HiSat2 and differential analysis was done in R Studio using the DESeq2 package. XGR, ExpressAnalyst and InnateDB algorithms were used for functional annotation and gene enrichment analysis of significant differentially expressed genes. RNA-seq was done on 23 r-HAT case samples and 28 healthy controls with 7 controls excluded for downstream analysis as outliers. A total of 4519 genes were significant differentially expressed (p adjusted <0.05) in individuals with early stage 1 r-HAT disease (n = 12) and 1824 genes in individuals with late stage 2 r-HAT disease (n = 11) compared to controls. Enrichment of innate immune response genes through neutrophil activation was identified in individuals with both early and late stages of the disease. Additionally, lipid metabolism genes were enriched in late stage 2 disease. We further identified uniquely upregulated genes (log2 Fold Change 1.4-2.0) in stage 1 (ZNF354C) and stage 2 (TCN1 and MAGI3) blood. Our data add to the current understanding of the human transcriptome profiles during T. b. rhodesiense infection. We further identified biological pathways and transcripts enriched than were enriched during stage 1 and stage 2 r-HAT. Lastly, we have identified transcripts which should be explored in future research whether they have potential of being used in combination with other markers for staging or r-HAT.

Transcriptome analysis of peripheral blood of Schistosoma mansoni infected children from the Albert Nile region in Uganda reveals genes implicated in fibrosis pathology.

Over 290 million people are infected by schistosomes worldwide. Schistosomiasis control efforts focus on mass drug treatment with praziquantel (PZQ), a drug that kills the adult worm of all Schistosoma species. Nonetheless, re-infections have continued to be detected in endemic areas with individuals living in the same area presenting with varying infection intensities. Our objective was to characterize the transcriptome profiles in peripheral blood of children between 10-15 years with varying intensities of Schistosoma mansoni infection living along the Albert Nile in Uganda. RNA extracted from peripheral blood collected from 44 S. mansoni infected (34 high and 10 low by circulating anodic antigen [CAA] level) and 20 uninfected children was sequenced using Illumina NovaSeq S4 and the reads aligned to the GRCh38 human genome. Differential gene expression analysis was done using DESeq2. Principal component analysis revealed clustering of gene expression by gender when S. mansoni infected children were compared with uninfected children. In addition, we identified 14 DEGs between S. mansoni infected and uninfected individuals, 56 DEGs between children with high infection intensity and uninfected individuals, 33 DEGs between those with high infection intensity and low infection intensity and no DEGs between those with low infection and uninfected individuals. We also observed upregulation and downregulation of some DEGs that are associated with fibrosis and its regulation. These data suggest expression of fibrosis associated genes as well as genes that regulate fibrosis in S. mansoni infection. The relatively few significant DEGS observed in children with schistosomiasis suggests that chronic S. mansoni infection is a stealth infection that does not stimulate a strong immune response.

Variants of IL6, IL10, FCN2, RNASE3, IL12B and IL17B loci are associated with Schistosoma mansoni worm burden in the Albert Nile region of Uganda.

BACKGROUND: Individuals genetically susceptible to high schistosomiasis worm burden may contribute disproportionately to transmission and could be prioritized for control. Identifying genes involved may guide development of therapy. METHODOLOGY/PRINCIPAL FINDINGS: A cohort of 606 children aged 10-15 years were recruited in the Albert Nile region of Uganda and assessed for Schistosoma mansoni worm burden using the Up-Converting Particle Lateral Flow (UCP-LF) test detecting circulating anodic antigen (CAA), point-of-care Circulating Cathodic Antigen (POC-CCA) and Kato-Katz tests. Whole genome genotyping was conducted on 326 children comprising the top and bottom 25% of worm burden. Linear models were fitted to identify variants associated with worm burden in preselected candidate genes. Expression quantitative trait locus (eQTL) analysis was conducted for candidate genes with UCP-LF worm burden included as a covariate. Single Nucleotide Polymorphism loci associated with UCP-LF CAA included IL6 rs2066992 (OR = 0.43, p = 0.0006) and rs7793163 (OR = 2.0, p = 0.0007); IL21 SNP kgp513476 (OR 1.79, p = 0.0025) and IL17B SNP kgp708159 (OR = 0.35, p = 0.0028). A haplotype in the IL10 locus was associated with lower worm burden (OR = 0.53, p = 0.015) and overlapped SNPs rs1800896, rs1800871 and rs1800872. Significant haplotypes (p<0.05, overlapping significant SNP) associated with worm burden were observed in IL6 and the Th17 pathway IL12B and IL17B genes. There were significant eQTL in the IL6, IL5, IL21, IL25 and IFNG regions. CONCLUSIONS: Variants associated with S. mansoni worm burden were in IL6, FCN2, RNASE3, IL10, IL12B and IL17B gene loci. However only eQTL associations remained significant after Bonferroni correction. In summary, immune balance, pathogen recognition and Th17 pathways may play a role in modulating Schistosoma worm burden. Individuals carrying risk variants may be targeted first in allocation of control efforts to reduce the burden of schistosomiasis in the community.

High prevalence of Schistosoma mansoni infection and stunting among school age children in communities along the Albert-Nile, Northern Uganda: A cross sectional study.

BACKGROUND: Knowing the prevalence of schistosomiasis is key to informing programmes to control and eliminate the disease as a public health problem. It is also important to understand the impact of infection on child growth and development in order to allocate appropriate resources and effort to the control of the disease. METHODS: We conducted a survey to estimate the prevalence of schistosomiasis among school aged children in villages along the Albert-Nile shore line in the district of Pakwach, North Western Uganda. A total of 914 children aged between 10-15 years were screened for Schistosoma mansoni using the POC-CCA and Kato Katz (KK) techniques. The infection intensities were assessed by POC-CCA and KK as well as CAA tests. The KK intensities were also correlated with POC-CCA and with CAA intensity. Anthropometric measurements were also taken and multivariate analysis was carried out to investigate their association with infection status. RESULTS: The prevalence of schistosomiasis using the POC-CCA diagnostic test was estimated at 85% (95% CI: 83-87), being highest amongst children living closer to the Albert-Nile shoreline. Visual scoring of the POC-CCA results was more sensitive than the Kato Katz test and was positively correlated with the quantified infection intensities by the CAA test. The majority of the children were underweight (BMI<18.5), and most notably, boys had significantly lower height for age (stunting) than girls in the same age range (p < 0.0001), but this was not directly associated with S. mansoni infection. CONCLUSION: High prevalence of S. mansoni infection in the region calls for more frequent mass drug administration with praziquantel. We observed high levels of stunting which was not associated with schistosomiasis. There is a need for improved nutrition among the children in the area.

In vitro culture of freshly isolated Trypanosoma brucei brucei bloodstream forms results in gene copy-number changes.

Most researchers who study unicellular eukaryotes work with an extremely limited number of laboratory-adapted isolates that were obtained from the field decades ago, but the effects of passage in laboratory rodents, and adaptation to in vitro culture, have been little studied. For example, the vast majority of studies of Trypanosoma brucei biology have concentrated on just two strains, Lister 427 and EATRO1125, which were taken from the field over half a century ago and have since have undergone innumerable passages in rodents and culture. We here describe two new Trypanosoma brucei brucei strains. MAK65 and MAK98, which have undergone only 3 rodent passages since isolation from Ugandan cattle. High-coverage sequencing revealed that adaptation of the parasites to culture was accompanied by changes in gene copy numbers. T. brucei has so far been considered to be uniformly diploid, but we also found trisomy of chromosome 5 not only in one Lister 427 culture, but also in the MAK98 field isolate. Trisomy of chromosome 6, and increased copies of other chromosome segments, were also seen in established cultured lines. The two new T. brucei strains should be useful to researchers interested in trypanosome differentiation and pathogenicity. Initial results suggested that the two strains have differing infection patterns in rodents. MAK65 is uniformly diploid and grew more reproducibly in bloodstream-form culture than MAK98.

Unmapped exome reads implicate a role for Anelloviridae in childhood HIV-1 long-term non-progression.

Human immunodeficiency virus (HIV) infection remains a significant public health burden globally. The role of viral co-infection in the rate of progression of HIV infection has been suggested but not empirically tested, particularly among children. We extracted and classified 42 viral species from whole-exome sequencing (WES) data of 813 HIV-infected children in Botswana and Uganda categorised as either long-term non-progressors (LTNPs) or rapid progressors (RPs). The Ugandan participants had a higher viral community diversity index compared to Batswana (p = 4.6 × 10-13), and viral sequences were more frequently detected among LTNPs than RPs (24% vs 16%; p = 0.008; OR, 1.9; 95% CI, 1.6-2.3), with Anelloviridae showing strong association with LTNP status (p = 3 × 10-4; q = 0.004, OR, 3.99; 95% CI, 1.74-10.25). This trend was still evident when stratified by country, sex, and sequencing platform, and after a logistic regression analysis adjusting for age, sex, country, and the sequencing platform (p = 0.02; q = 0.03; OR, 7.3; 95% CI, 1.6-40.5). Torque teno virus (TTV), which made up 95% of the Anelloviridae reads, has been associated with reduced immune activation. We identify an association between viral co-infection and prolonged AIDs-free survival status that may have utility as a biomarker of LTNP and could provide mechanistic insights to HIV progression in children, demonstrating the added value of interrogating off-target WES reads in cohort studies.

Molecular characterization of non-polio enteroviruses isolated from acute flaccid paralysis patients in Uganda.

Enteroviruses (EVs) are RNA viruses that can cause many clinical syndromes including acute flaccid paralysis (AFP). Within the global polio laboratory network, EVs are categorized either as polioviruses or non-polio enteroviruses (NPEVs). Specific NPEVs have been described in polio-like residual paralytic events in AFP patients. Retrospective analysis of 112 NPEV isolates from AFP patients was performed and thirty one NPEV types were identified of which 91% were Enterovirus B and 9% were Enterovirus A species. The NPEVs were distributed across the country with most patients in the eastern region (41/89; 46.1%). The highest proportion of patients were children less than 5 years (77/89; 86.5%) and male patients were more common (54/89; 60.7%). Echovirus 11 (11/89; 12.4%) was frequently observed and phylogenetic analysis of these sequences revealed high diversity. Coxsackievirus B5 (CV-B5), CV-B6, E21, and EV-B69 were only seen in patients with residual paralysis. Analyses of the EV-A71 sequence indicated a unique genogroup.

Candidate gene family-based and case-control studies of susceptibility to high Schistosoma mansoni worm burden in African children: a protocol.

Background: Approximately 25% of the risk of Schistosoma mansoni is associated with host genetic variation. We will test 24 candidate genes, mainly in the T h2 and T h17 pathways, for association with S. mansoni infection intensity in four African countries, using family based and case-control approaches. Methods: Children aged 5-15 years will be recruited in S. mansoni endemic areas of Ivory Coast, Cameroon, Uganda and the Democratic Republic of Congo (DRC). We will use family based (study 1) and case-control (study 2) designs. Study 1 will take place in Ivory Coast, Cameroon, Uganda and the DRC. We aim to recruit 100 high worm burden families from each country except Uganda, where a previous study recruited at least 40 families. For phenotyping, cases will be defined as the 20% of children in each community with heaviest worm burdens as measured by the circulating cathodic antigen (CCA) assay. Study 2 will take place in Uganda. We will recruit 500 children in a highly endemic community. For phenotyping, cases will be defined as the 20% of children with heaviest worm burdens as measured by the CAA assay, while controls will be the 20% of infected children with the lightest worm burdens. Deoxyribonucleic acid (DNA) will be genotyped on the Illumina H3Africa SNP (single nucleotide polymorphisms) chip and genotypes will be converted to sets of haplotypes that span the gene region for analysis. We have selected 24 genes for genotyping that are mainly in the Th2 and Th17 pathways and that have variants that have been demonstrated to be or could be associated with Schistosoma infection intensity.   Analysis: In the family-based design, we will identify SNP haplotypes disproportionately transmitted to children with high worm burden. Case-control analysis will detect overrepresentation of haplotypes in extreme phenotypes with correction for relatedness by using whole genome principal components.

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.

High-Throughput Sequencing for Trypanosome Transcriptome Characterization.

High-throughput sequencing of cDNA (RNASeq) is now the method of choice for analysis of transcriptomes. This chapter details important considerations in the design of RNASeq experiments for kinetoplastids grown in culture or experimental animals. It contains protocols for obtaining parasites from rodents, and for removal of rRNA from total RNA. In addition, custom pipelines for sequence alignment, and for data analysis and visualization, are described.

Blood signatures for second stage human African trypanosomiasis: a transcriptomic approach.

BACKGROUND: Rhodesiense sleeping sickness is caused by infection with T. b rhodesiense parasites resulting in an acute disease that is fatal if not treated in time. The aim of this study was to understand the global impact of active T. b rhodesiense infection on the patient's immune response in the early and late stages of the disease. METHODS: RNASeq was carried out on blood and cerebral spinal fluid (CSF) samples obtained from T. b. rhodesiense infected patients. The control samples used were from healthy individuals in the same foci. The Illumina sequenced reads were analysed using the Tuxedo suite pipeline (Tophat, Cufflinks, Cuffmerge, Cuffdiff) and differential expression analysis carried out using the R package DESeq2. The gene enrichment and function annotation analysis were done using the ToppCluster, DAVID and InnateDB algorithms. RESULTS: We previously described the transcriptomes of T. b rhodesiense from infected early stage blood (n = 3) and late stage CSF (n = 3) samples from Eastern Uganda. We here identify human transcripts that were differentially expressed (padj < 0.05) in the early stage blood versus healthy controls (n = 3) and early stage blood versus late stage CSF. Differential expression in infected blood showed an enrichment of innate immune response genes whereas that of the CSF showed enrichment for anti-inflammatory and neuro-degeneration signalling pathways. We also identified genes (C1QC, MARCO, IGHD3-10) that were up-regulated (log2 FC > 2.5) in both the blood and CSF. CONCLUSION: The data yields insights into the host's response to T. b rhodesiense parasites in the blood and central nervous system. We identified key pathways and signalling molecules for the predominant innate immune response in the early stage infection; and anti-inflammatory and neuro-degeneration pathways associated with sleep disorders in second stage infection. We further identified potential blood biomarkers that can be used for diagnosis of late stage disease without the need for lumbar puncture.

De Novo Genome Sequence of a Fusarium xylarioides Race Pathogenic to Robusta Coffee (Coffea canephora) in Uganda.

Here, we report the first whole-genome assembly of a Fusarium xylarioides race pathogenic to robusta coffee in Uganda. It comprises 55,122,624 bases and 14,552 genes. Gene ontology analysis assigned 5,720 genes to biological processes, 4,545 genes to cellular components, and 6,021 genes to molecular function.

Transcriptomes of Trypanosoma brucei rhodesiense from sleeping sickness patients, rodents and culture: Effects of strain, growth conditions and RNA preparation methods.

All of our current knowledge of African trypanosome metabolism is based on results from trypanosomes grown in culture or in rodents. Drugs against sleeping sickness must however treat trypanosomes in humans. We here compare the transcriptomes of Trypanosoma brucei rhodesiense from the blood and cerebrospinal fluid of human patients with those of trypanosomes from culture and rodents. The data were aligned and analysed using new user-friendly applications designed for Kinetoplastid RNA-Seq data. The transcriptomes of trypanosomes from human blood and cerebrospinal fluid did not predict major metabolic differences that might affect drug susceptibility. Usefully, there were relatively few differences between the transcriptomes of trypanosomes from patients and those of similar trypanosomes grown in rats. Transcriptomes of monomorphic laboratory-adapted parasites grown in in vitro culture closely resembled those of the human parasites, but some differences were seen. In poly(A)-selected mRNA transcriptomes, mRNAs encoding some protein kinases and RNA-binding proteins were under-represented relative to mRNA that had not been poly(A) selected; further investigation revealed that the selection tends to result in loss of longer mRNAs.

No evidence for association between APOL1 kidney disease risk alleles and Human African Trypanosomiasis in two Ugandan populations.

BACKGROUND: Human African trypanosomiasis (HAT) manifests as an acute form caused by Trypanosoma brucei rhodesiense (Tbr) and a chronic form caused by Trypanosoma brucei gambiense (Tbg). Previous studies have suggested a host genetic role in infection outcomes, particularly for APOL1. We have undertaken candidate gene association studies (CGAS) in a Ugandan Tbr and a Tbg HAT endemic area, to determine whether polymorphisms in IL10, IL8, IL4, HLAG, TNFA, TNX4LB, IL6, IFNG, MIF, APOL1, HLAA, IL1B, IL4R, IL12B, IL12R, HP, HPR, and CFH have a role in HAT. METHODOLOGY AND RESULTS: We included 238 and 202 participants from the Busoga Tbr and Northwest Uganda Tbg endemic areas respectively. Single Nucleotide Polymorphism (SNP) genotype data were analysed in the CGAS. The study was powered to find odds ratios > 2 but association testing of the SNPs with HAT yielded no positive associations i.e. none significant after correction for multiple testing. However there was strong evidence for no association with Tbr HAT and APOL1 G2 of the size previously reported in the Kabermaido district of Uganda. CONCLUSIONS/SIGNIFICANCE: A recent study in the Soroti and Kaberamaido focus in Central Uganda found that the APOL1 G2 allele was strongly associated with protection against Tbr HAT (odds ratio = 0.2, 95% CI: 0.07 to 0.48, p = 0.0001). However, in our study no effect of G2 on Tbr HAT was found, despite being well powered to find a similar sized effect (OR = 0.9281, 95% CI: 0.482 to 1.788, p = 0.8035). It is possible that the G2 allele is protective from Tbr in the Soroti/Kabermaido focus but not in the Iganga district of Busoga, which differ in ethnicity and infection history. Mechanisms underlying HAT infection outcome and virulence are complex and might differ between populations, and likely involve several host, parasite or even environmental factors.

Candidate genes-based investigation of susceptibility to Human African Trypanosomiasis in Côte d'Ivoire.

Human African Trypanosomiasis (HAT) or sleeping sickness is a Neglected Tropical Disease. Long regarded as an invariably fatal disease, there is increasing evidence that infection by T. b. gambiense can result in a wide range of clinical outcomes, including latent infections, which are long lasting infections with no parasites detectable by microscopy. The determinants of this clinical diversity are not well understood but could be due in part to parasite or host genetic diversity in multiple genes, or their interactions. A candidate gene association study was conducted in Côte d'Ivoire using a case-control design which included a total of 233 subjects (100 active HAT cases, 100 controls and 33 latent infections). All three possible pairwise comparisons between the three phenotypes were tested using 96 SNPs in16 candidate genes (IL1, IL4, IL4R, IL6, IL8, IL10, IL12, IL12R, TNFA, INFG, MIF, APOL1, HPR, CFH, HLA-A and HLA-G). Data from 77 SNPs passed quality control. There were suggestive associations at three loci in IL6 and TNFA in the comparison between active cases and controls, one SNP in each of APOL1, MIF and IL6 in the comparison between latent infections and active cases and seven SNP in IL4, HLA-G and TNFA between latent infections and controls. No associations remained significant after Bonferroni correction, but the Benjamini Hochberg false discovery rate test indicated that there were strong probabilities that at least some of the associations were genuine. The excess of associations with latent infections despite the small number of samples available suggests that these subjects form a distinct genetic cluster different from active HAT cases and controls, although no clustering by phenotype was observed by principle component analysis. This underlines the complexity of the interactions existing between host genetic polymorphisms and parasite diversity.

A polymorphism in the haptoglobin, haptoglobin related protein locus is associated with risk of human sleeping sickness within Cameroonian populations.

BACKGROUND: Human African Trypanosomiasis (HAT) is a neglected disease targeted for elimination as a public health problem by 2020. Elimination requires a better understanding of the epidemiology and clinical evolution of HAT. In addition to the classical clinical evolution of HAT, asymptomatic carriers and spontaneous cure have been reported in West Africa. A genetic component to human susceptibility to HAT has been suggested to explain these newly observed responses to infection. In order to test for genetic associations with infection response, genetic polymorphism in 17 genes were tested (APOL1, IL1B, IL4, IL4R, IL6, IL8, IL12B, IL12RB1, IL10, TNFA, INFG, MIF, HLA-G, HLA-A, HP, HPR and CFH). METHODOLOGY: A case-control study was performed on 180 blood samples collected from 56 cases and 124 controls from Cameroon. DNA was extracted from blood samples. After quality control, 25 samples (24 controls and 1 case) were eliminated. The genotyping undertaken on 155 individuals including 55 cases and 100 controls were investigated at 96 loci (88 SNPs and 8 indels) located on 17 genes. Associations between these loci and HAT were estimated via a case-control association test. RESULTS: Analyses of 64 SNPs and 4 indels out of 96 identified in the selected genes reveal that the minor allele (T) of rs8062041 in haptoglobin (HP) appeared to be protective against HAT (p = 0.0002395, OR 0.359 (CI95 [0.204-0.6319])); indicating higher frequency in cases compared to controls. This minor allele with adjusted p value of 0.0163 is associated with a lower risk (protective effect) of developing sleeping sickness. CONCLUSION: The haptoglobin related protein HPR and HP are tightly linked and both are duplicated in some people and may lead to higher activity. This increased production could be responsible of the protection associated with rs8062041 even though this SNP is within HP.

Prevalence of pathogenic free-living amoeba and other protozoa in natural and communal piped tap water from Queen Elizabeth protected area, Uganda.

BACKGROUND: Pathogenic water dwelling protozoa such as Acanthamoeba spp., Hartmannella spp., Naegleria spp., Cryptosporidium spp. and Giardia spp. are often responsible for devastating illnesses especially in children and immunocompromised individuals, yet their presence and prevalence in certain environment in sub-Saharan Africa is still unknown to most researchers, public health officials and medical practitioners. The objective of this study was to establish the presence and prevalence of pathogenic free-living amoeba (FLA), Cryptosporidium and Giardia in Queen Elizabeth Protected Area (QEPA). METHODS: Samples were collected from communal taps and natural water sites in QEPA. Physical water parameters were measured in situ. The samples were processed to detect the presence of FLA trophozoites by xenic cultivation, Cryptosporidium oocysts by Ziehl-Neelsen stain and Giardia cysts by Zinc Sulphate floatation technique. Parasites were observed microscopically, identified, counted and recorded. For FLA, genomic DNA was extracted for amplification and sequencing. RESULTS: Both natural and tap water sources were contaminated with FLA, Cryptosporidium spp. and Giardia spp. All protozoan parasites were more abundant in the colder rainy season except for Harmannella spp. and Naegleria spp. which occurred more in the warmer months. The prevalence of all parasites was higher in tap water than in natural water samples. There was a strong negative correlation between the presence of Acanthamoeba spp., Hartmannella spp., Cryptosporidium spp. and Giardia spp. with Dissolved Oxygen (DO) (P < 0.05). The presence of Cryptosporidium spp. showed a significant positive correlation (P < 0.05) with conductivity, pH and Total Dissolved Solids (TDS); whereas the presence of Giardia spp. had only a strong positive correlation with TDS. Molecular genotyping of FLA produced 7 Acanthamoeba, 5 Echinamoeba, 2 Hartmannella, 1 Bodomorpha, 1 Nuclearia and 1 Cercomonas partial sequences. CONCLUSIONS: All water collection sites were found to be contaminated with pathogenic protozoa that could possibly be the cause of a number of silent morbidities and mortalities among rural households in QEPA. This implies that water used by communities in QEPA is of poor quality and predisposes them to a variety of protozoan infections including the FLA whose public health importance was never reported, thus necessitating adoption of proper water safety measures.