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Every novel infection requires an assessment of the host response coupled with identification of unique biomarkers for predicting disease pathogenesis, treatment targets and diagnostic utility. Studies have exposed dysregulated inflammatory response induced by the novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) as significant predictor or cause of disease severity/prognosis and death. This study evaluated inflammatory biomarkers induced by SARS-CoV-2 in plasma of patients with varying disease phenotypes and healthy controls with prognostic or therapeutic potential. We stratified SARS-CoV-2 plasma samples based on disease status (asymptomatic, mild, severe, and healthy controls), as diagnosed by RT-PCR SARS-CoV-2. We used a solid phase sandwich and competitive Enzyme-Linked Immunosorbent Assay (ELISA) to measure levels of panels of immunological (IFN-γ, TNF-α, IL-6, and IL-10) and biochemical markers (Ferritin, Procalcitonin, C-Reactive Protein, Angiotensin II, Homocysteine, and D-dimer). Biomarker levels were compared across SARS-CoV-2 disease stratification. Plasma IFN-γ, TNF-α, IL-6, and IL-10 levels were significantly (P < 0.05) elevated in the severe SARS-CoV-2 patients as compared to mild, asymptomatic, and healthy controls. Ferritin, Homocysteine, and D-dimer plasma levels were significantly elevated in severe cases over asymptomatic and healthy controls. Plasma C-reactive protein and Angiotensin II levels were significantly (P < 0.05) higher in mild than severe cases and healthy controls. Plasma Procalcitonin levels were significantly higher in asymptomatic than in mild, severe cases and healthy controls. Our study demonstrates the role of host inflammatory biomarkers in modulating the pathogenesis of COVID-19. The study proposes a number of potential biomarkers that could be explored as SARS-CoV-2 treatment targets and possible prognostic predictors for a severe outcome. The comprehensive analysis of prognostic biomarkers may contribute to the evidence-based management of COVID-19 patients. [ABSTRACT FROM AUTHOR]

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. Author summary: This study investigated genetic risk factors for high worm burden in a cohort of 606 children aged 10–15 years old in the Albert Nile region of Uganda. Genotyping was conducted on 326 children who comprised the top and bottom 25% of worm burden. Results showed single nucleotide polymorphisms (SNPs) associated with worm burden in the IL6, IL21 and IL17B genes, and a haplotype in the IL10 locus associated with lower worm burden. Expression quantitative trait locus (eQTL) analysis revealed significant gene and worm burden associations in the IL6 region, and expression associations in the IL5, IL21, IL25 and IFNG loci. These findings suggest that immune balance, pathogen recognition and Th17 pathways may be involved in modulating Schistosoma worm burden and that individuals carrying risk variants may be prioritized to improve control efforts. [ABSTRACT FROM AUTHOR]

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. Author summary: Schistosomiasis is a neglected tropical disease transmitted via an intermediate snail host through contact with contaminated fresh water. Even with routine Mass Drug Administration for treatment of the infection, re-infections are still common and variations in infection intensity and pathology are still observed in individuals in the same location. These may be due to differences in individuals' response to S. mansoni infection. In this study, we used RNAseq to identify differentially expressed genes associated with S. mansoni infection in children between 10–15 years. We conducted comparisons between phenotypes including infection intensities measured by circulating anodic antigen, wasting by body mass index and stunting by height-for-age Z-score. Our data showed very low numbers of significantly differentially expressed genes in all comparisons. Some of the few differentially expressed genes that were observed were associated with fibrosis which is the cause of pathology in humans and has been observed in late stages of S. mansoni infection in murine studies. [ABSTRACT FROM AUTHOR]

The integrated control of East Coast fever (ECF) by early diagnosis and treatment involving acquired immunity induced by natural infection in Ankole cattle was assessed. A longitudinal study was carried out in Kiruhura district, southwestern Uganda for six months on 244 Ankole breed of cattle from 18 herds under natural tick challenge and relaxed tick control measures. Calves aged three to six months old were recruited and monitored daily by farmers for detection of ECF clinical symptoms. The reported sick animals were treated using Buparvaquone and treatment outcome determined. Monthly follow-ups and blood collections were done to monitor ECF status. Blood was analyzed for Theileria parva parasites by microscopy, DNA by polymerase chain reaction (PCR) and antibodies by enzyme-linked immunosorbent assay (ELISA). The overall prevalence of ECF clinical disease within six months period was 30.3% (74). The major symptoms of early clinical ECF disease were fever and enlarged parotid or prescapular lymph nodes. Clinical cases were categorized as mild, 24% (18) or moderate, 76% (56). There was an overall recovery rate of 100% (74) of the ECF cases whereby 94.6% (70) recovered promptly and 5.4% (4) recovered slowly. Based on blood analysis, prevalence of ECF at baseline was 3.7% (9) by microscopy, 31.1% (76) by PCR and 38.1% (93) by ELISA. A significant increase (p < 0.05) was shown by the increased number of calves with T. parva specific antibodies in the sera from 38.1% at baseline to 68.8% after six months. High antibody levels (positive percentage ≥ 50%) were detected in all ECF-treated and recovered calves at the end of six months. The acquired immunity to ECF was high in treated and recovered cattle, indicating that natural exposure to infection, accurate early diagnosis and effective treatment enhance development of immune-protection in indigenous cattle in an endemic area. The prominent early clinical symptoms for ECF could be exploited in the development of decision support tools for chemotherapy and other integrated control measures. [ABSTRACT FROM AUTHOR]

Background: Malaria is endemic in all regions where gambiense or rhodesiense human African trypanosomiasis (HAT) is reported, and both diseases have similarities in their symptomatology. A combined test could be useful for both diseases and would facilitate integration of the screening for gambiense HAT (gHAT) and malaria diagnosis. This study aimed to evaluate a combined prototype rapid diagnostic test (RDT) for gHAT and malaria. Methods: Blood samples were collected in the Democratic Republic of the Congo and in Uganda to evaluate the performance of a prototype HAT/Malaria Combined RDT in comparison to an individual malaria RDT based on Plasmodium falciparum (P.f.) Histidine Rich Protein II (HRP-II or HRP2) antigen (SD BIOLINE Malaria Ag P.f. RDT) for malaria detection and an individual gHAT RDT based on recombinant antigens, the SD BIOLINE HAT 2.0 RDT for HAT screening. Due to the current low prevalence of gHAT in endemic regions, the set of blood samples that were collected was used to evaluate the specificity of the RDTs for gHAT, and additional archived plasma samples were used to complete the evaluation of the HAT/Malaria Combined RDT in comparison to the HAT 2.0 RDT. Results: Frozen whole blood samples from a total of 486 malaria cases and 239 non-malaria controls, as well as archived plasma samples from 246 gHAT positive and 246 gHAT negative individuals were tested. For malaria, the sensitivity and specificity of the malaria band in the HAT/Malaria Combined RDT were 96.9% (95% CI: 95.0–98.3) and 97.1% (95% CI: 94.1–98.8) respectively. The sensitivity and specificity of the SD BIOLINE malaria Ag P.f. RDT were 97.3% (95% CI: 95.5–98.6) and 97.1% (95% CI: 94.1–98.8) respectively. For gHAT, using archived plasma samples, the sensitivity and specificity were respectively 89% (95% CI: 84.4–92.6) and 93.5% (95% CI: 89.7–96.2) with the HAT /malaria Combined RDT, and 88.2% (95% CI: 83.5–92) and 94.7% (95% CI: 91.1–97.2) with the HAT 2.0 RDT. Using the whole blood samples that were collected during the study, the specificity of the HAT/malaria Combined RDT for gHAT was 95.8% (95% CI: 94.3–97.0). Conclusion: The HAT/Malaria Combined prototype RDT was as accurate as the individual malaria or gHAT RDTs. The HAT/Malaria Combined prototype RDT is therefore suitable for both malaria diagnosis and gHAT screening. However, there is a need to assess its accuracy using fresh samples in prospective clinical trials. Author summary: The annual number of reported cases of human African trypanosomiasis (HAT), also known as sleeping sickness (SS), is currently below 1,000 cases worldwide. The Democratic Republic of the Congo (DRC), the most affected country, and Uganda, which shares a border with DRC, are both endemic for gambiense HAT (gHAT). The main strategy to control gHAT is active screening of at-risk individuals, followed by diagnosis and treatment of confirmed cases. However, this strategy and even the passive screening as currently implemented become less efficient with declining incidence, justifying innovative strategies to efficiently detect the remaining cases. All areas where gHAT occurs are also endemic for malaria, presenting an opportunity to integrate gHAT screening activities within malaria control activities. This integration is warranted by the fact that in early disease stage, gHAT patients present with signs and symptoms strikingly similar to those of malaria. In order to use malaria diagnosis as an entry point to screen for gHAT, Standard Diagnostics (SD), Korea (now Abbott Diagnostics, Korea Inc–ADK) made a Combined prototype RDT for both malaria and gHAT, expected to be as accurate as the individual gHAT and malaria RDTs. In this study, we evaluated the accuracy of the Combined prototype RDT using whole blood samples collected in Uganda and DRC, and archived plasma samples collected in DRC, Angola and Central African Republic. We found that the Combined prototype performs just as well as individual RDTs. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Background: Immunological Human African Trypanosomiasis (HAT) studies often exclude malaria, although both infections overlap in specific endemic areas. During this co-infection, it is not known whether this parasitic interaction induces synergistic or antagonistic cytokine response among humans. This study determined prevalence of Plasmodium falciparum malaria among Trypanosoma brucei rhodesiense HAT and plasma cytokine profile levels associated with HAT and/or malaria infections. Methods: Participants were recruited at Lwala hospital in north eastern Uganda: healthy controls (30), malaria (28), HAT (17), HAT and malaria (15) diagnosed by microscopy and PCR was carried out for parasite species identification. Plasma cytokine levels of Interferon-gamma (IFN-γ), Tumour Necrosis Factor-alpha (TNF-α), Interleukin (IL)-6, IL-10 and Transforming Growth Factor-beta (TGF-β) were measured by sandwich Enzyme-Linked Immuno Sorbent Assay and data statistically analysed using Graphpad Prism 6.0. Results: The prevalence of P. falciparum malaria among T. rhodesiense HAT cases was high (46.8%). Malaria and/or HAT cases presented significant higher plasma cytokine levels of IFN-γ, TNF-α, IL-6, IL-10 and TGF-β than healthy controls (P < 0.05). Levels of IFN-γ, IL-6 and IL-10 were significantly elevated in HAT over malaria (P < 0.05) but no significant difference in TNF-α and TGF-β between HAT and malaria (P > 0.05). Co-infection expressed significantly higher plasma IFN-γ, IL-6, and IL-10 levels than malaria (P < 0.05) but no significant difference with HAT mono-infection (P > 0.05). The TNF-α level was significantly elevated in co-infection over HAT or malaria mono-infections (P < 0.05) unlike TGF-β level. Significant positive correlations were identified between IFN-γ verses TNF-α and IL-6 verses IL-10 in co-infection (Spearman's P < 0.05). Conclusions: The T. b. rhodesiense significantly induced the cytokine response more than P. falciparum infections. Co-infection led to synergistic stimulation of pro-inflammatory (IFN-γ, TNF-α), and anti-inflammatory (IL-6, and IL-10) cytokine responses relative to malaria mono-infection. Level of TNF-α partially indicates the effect induced by T. b. rhodesiense and P. falciparum mono-infections or a synergistic interaction of co-infections which may have adverse effects on pathogenesis, prognosis and resolution of the infections. Trial registration VCD-IRC/021, 26/08/2011; HS 1089, 16/01/2012 [ABSTRACT FROM AUTHOR]

Trypanosoma brucei (T.b.) rhodesiense is the cause of the acute form of human African trypanosomiasis (HAT) in eastern and southern African countries. There is some evidence that there is diversity in the disease progression of T.b. rhodesiense in different countries. HAT in Malawi is associated with a chronic haemo-lymphatic stage infection compared to other countries, such as Uganda, where the disease is acute with more marked neurological impairment. This has raised the question of the role of host genetic factors in infection outcomes. A candidate gene association study was conducted in the northern region of Malawi. This was a case-control study involving 202 subjects, 70 cases and 132 controls. All individuals were from one area; born in the area and had been exposed to the risk of infection since birth. Ninety-six markers were genotyped from 17 genes: IL10, IL8, IL4, HLA-G, TNFA, IL6, IFNG, MIF, APOL, HLA-A, IL1B, IL4R, IL12B, IL12R, HP, HPR, and CFH. There was a strong significant association with APOL1 G2 allele (p = 0.0000105, OR = 0.14, CI95 = [0.05–0.41], BONF = 0.00068) indicating that carriers of the G2 allele were protected against T.b. rhodesiense HAT. SNP rs2069845 in IL6 had raw p < 0.05, but did not remain significant after Bonferroni correction. There were no associations found with the other 15 candidate genes. Our finding confirms results from other studies that the G2 variant of APOL1 is associated with protection against T.b. rhodesiense HAT. [ABSTRACT FROM AUTHOR]

Introduction: The incidence of gambiense human African trypanosomiasis (gHAT) in Uganda has been declining, from 198 cases in 2008, to only 20 in 2012. Interruption of transmission of the disease by early diagnosis and treatment is core to the control and eventual elimination of gHAT. Until recently, the format of available screening tests had restricted screening and diagnosis to central health facilities (passive screening). We describe a novel strategy that is contributing to elimination of gHAT in Uganda through expansion of passive screening to the entire population at risk. Methodology / Principal findings: In this strategy, patients who are clinically suspected of having gHAT at primary health facilities are screened using a rapid diagnostic test (RDT), followed by parasitological confirmation at strategically located microscopy centres. For patients who are positive with the RDT and negative by microscopy, blood samples undergo further testing using loop-mediated isothermal amplification (LAMP), a molecular test that detects parasite DNA. LAMP positive patients are considered strong suspects, and are re-evaluated by microscopy. Location and upgrading of facilities to perform microscopy and LAMP was informed by results of georeferencing and characterization of all public healthcare facilities in the 7 gHAT endemic districts in Uganda. Three facilities were upgraded to perform RDTs, microscopy and LAMP, 9 to perform RDTs and microscopy, and 200 to screen patients with RDTs. This reduced the distance that a sick person must travel to be screened for gHAT to a median distance of 2.5km compared to 23km previously. In this strategy, 9 gHAT cases were diagnosed in 2014, and 4 in 2015. Conclusions: This enhanced passive screening strategy for gHAT has enabled full coverage of the population at risk, and is being replicated in other gHAT endemic countries. The improvement in case detection is making elimination of the disease in Uganda an imminent possibility. [ABSTRACT FROM AUTHOR]

Background: Brucellosis remains a neglected zoonotic disease among agro-pastoral communities where unprocessed milk and milk products are consumed. A cross-sectional study was carried out in Kiboga district to determine the seroprevalence and risk factors associated with human brucellosis in communities where livestock rearing in a common practice.Methods: A total of 235 participants were involved in the study. Blood samples from the participants were collected and screened for Brucella using Serum Agglutination Test and Rose Bengal Plate Test. A questionnaire was used to collect data on socio-demographic characteristics and human brucellosis related risk factors.Results: Human Brucella seroprevalence was at 17.0 % (n = 235). The prevalence was highest among males (20.5 %, n = 78) and the elderly - above 60 years (22.2 %, n = 18). Residence in rural areas (OR 3.16, 95 % CI: 1.16-8.56), consuming locally processed milk products (OR 2.54, 95 % CI: 1.12-5.78) and being single (OR 2.44, 95 % CI: 1.05-5.68), were associated with increased risk of brucellosis.Discussion: Human brucellosis seroprevalence was high at 17 %, this was parallel with animal brucellosis prevalence that has been reported to range from 10.2 % to 25.7 % in cattle in the region. The participants were from communities known to habitually consume raw milk and milk products, know to process milk products using bare hands which are major risk factors for brucellosis in humans. This also explains why consumption of unpasteurized milk products was associated with the occurrence of brucellosis in study area. This strengthened the argument that humans get infected through consumption of contaminated animal products as reported in other earlier studies. Males and elderly being more affected because of traditional roles of these groups they play in livestock care and management. The single were also to be more associated to brucellosis, due to the fact that this group consume milk and milk products more as it is readily available in the informal markets as highly nutritious fast foods in this community as also observed in USA.Conclusions: Brucellosis is highly prevalent in Kiboga district, and therefore, an important public health problem. The transmission risk was aggravated by consumption of unpasteurized milk products, residing in rural settings and being single. There is a need to initiate screening, treat infected humans early, and educate the public about risk factors and appropriate preventive measures of brucellosis. [ABSTRACT FROM AUTHOR]

Background: Sleeping sickness due to Trypanosoma brucei rhodesiense has a wide spectrum of clinical presentations coupled with differences in disease progression and severity across East and Southern Africa. The disease progresses from an early (hemo-lymphatic) stage to the late (meningoencephalitic) stage characterized by presence of parasites in the central nervous system. We hypothesized that disease progression and severity of the neurological response is modulated by cytokines. Methods: A total of 55 sleeping sickness cases and 41 healthy controls were recruited passively at Lwala hospital, in Northern Uganda. A panel of six cytokines (IFN-γ, IL1-β, TNF-α, IL-6, TGF-β and IL-10) were assayed from paired plasma and cerebrospinal fluid (CSF) samples. Cytokine concentrations were analyzed in relation to disease progression, clinical presentation and severity of neurological responses. Results: Median plasma levels (pg/ml) of IFN-γ (46.3), IL-6 (61.7), TGF-β (8755) and IL-10 (256.6) were significantly higher in cases compared to controls (p< 0.0001). When early stage and late stage CSF cytokines were compared, IL-10 and IL-6 were up regulated in late stage patients and were associated with a reduction in tremors and cranioneuropathy. IL-10 had a higher staging accuracy with a sensitivity of 85.7% (95% CI, 63.7%-97%) and a specificity of 100% (95% CI, 39.8%-100%) while for IL-6, a specificity of 100% (95% CI, 47.8%-100%) gave a sensitivity of 83.3% (95% CI, 62.2%-95.3%). Conclusion: Our study demonstrates the role of host inflammatory cytokines in modulating the progression and severity of neurological responses in sleeping sickness. We demonstrate here an up-regulation of IL-6 and IL-10 during the late stage with a potential as adjunct stage biomarkers. Given that both cytokines could potentially be elevated by other CNS infections, our findings should be further validated in a large cohort of patients including those with other inflammatory diseases such as cerebral malaria. Author Summary: Sleeping sickness in east and central Africa is caused by a protozoan parasite, Trypanosoma brucei rhodesiense. About 12.3 million people are at a risk of acquiring the disease that is fatal if untreated. The disease progresses from an early stage with trypanosomes in blood and lymph to a late stage in which trypanosomes enter the central nervous system. Variations in disease presentation, progression and severity of the disease have been reported and cytokines have been proposed as potential players. Before treatment is commenced, disease stage has to be ascertained since treatment for the two stages is different. The currently used staging criteria depends on finding trypanosomes in the cerebrospinal fluids (CSF) and elevation in CSF white blood cells however, this has been found to have low sensitivity. We therefore measured plasma and cerebrospinal fluid cytokine concentrations and determined associations with disease presentation, stage progression and severity of the neurological response. Our study shows that concentrations of specific cytokines are elevated in sleeping sickness patients and have a potential to discriminate between patients in early or late stage of infection. [ABSTRACT FROM AUTHOR]

Background: The acute form of Human African Trypanosomiasis (HAT, also known as Sleeping sickness) caused by Trypanosoma brucei rhodesiense has been shown to have a wide spectrum of focus specific clinical presentation and severity in East and Southern Africa. Indeed HAT occurs in regions endemic for other tropical diseases, however data on how these co-morbidities might complicate the clinical picture and affect disease outcome remains largely scanty. We here describe the clinical presentation, presence of co-infections, and how the latter impact on HAT prognosis. Methods and Findings: We carried out a retrospective analysis of clinical data from 258 sleeping sickness patients reporting to Lwala hospital between 2005 and 2012. The mean patient age was 28.6 years with a significant number of cases below 18 years (p< 0.0001). About 93.4% of the cases were diagnosed as late stage (p< 0.0001). The case fatality rate was 10.5% with post treatment reactive encephalopathys reported in 7.9% of the cases, of whom 36.8% eventually died. Fever was significantly (p = 0.045) higher in patients under 18 years. Of the early stage patients, 26.7% and 6.7% presented with late stage signs of sleep disorder and mental confusion respectively. Among the co-infections, malaria was significantly more prevalent (28.9%; p< 0.0001) followed by urinary tract infections (4.2%). Co-infections were present in 14.3% of in-hospital deaths, 38.5% of which were recorded as Malaria. Malaria was significantly more common in patients under 18 years (45.5%; p< 0.02), and was reported in 60% of the fatal cases in this age group. Conclusions: We show a wide spectrum of sleeping sickness clinical presentation and disease outcome that was apparently not significantly influenced by concurrent infections. It would thus be interesting to determine the host and/or parasite factors that might be responsible for the observed diverse clinical presentation. [ABSTRACT FROM AUTHOR]

Background: The polymerase chain reaction (PCR) and nucleic acid sequence-based amplification (NASBA) have been recently modified by coupling to oligochromatography (OC) for easy and fast visualisation of products. In this study we evaluate the sensitivity and specificity of the PCR-OC and NASBA-OC for diagnosis of Trypanosoma brucei gambiense and Trypanosoma brucei rhodesiense human African trypanosomiasis (HAT). Methodology and Results: Both tests were evaluated in a case-control design on 143 HAT patients and 187 endemic controls from the Democratic Republic of Congo (DRC) and Uganda. The overall sensitivity of PCR-OC was 81.8% and the specificity was 96.8%. The PCR-OC showed a sensitivity and specificity of 82.4% and 99.2% on the specimens from DRC and 81.3% and 92.3% on those from Uganda. NASBA-OC yielded an overall sensitivity of 90.2%, and a specificity of 98.9%. The sensitivity and specificity of NASBA-OC on the specimens from DRC was 97.1% and 99.2%, respectively. On the specimens from Uganda we observed a sensitivity of 84.0% and a specificity of 98.5%. Conclusions/Significance: The tests showed good sensitivity and specificity for the T. b. gambiense HAT in DRC but rather a low sensitivity for T. b. rhodesiense HAT in Uganda. Author Summary: Diagnosis plays a central role in the control of human African trypanosomiasis (HAT) whose mainstay in disease control is chemotherapy. However, accurate diagnosis is hampered by the absence of sensitive techniques for parasite detection. Without concentrating the blood, detection thresholds can be as high as 10,000 trypanosomes per milliliter of blood. The polymerase chain reaction (PCR) and nucleic acid sequence-based amplification (NASBA) are promising molecular diagnostics that generally yield high sensitivity and could improve case detection. Recently, these two tests were coupled to oligochromatography (OC) for simplified and standardized detection of amplified products, eliminating the need for electrophoresis. In this study, we evaluated the diagnostic accuracy of these two novel tests on blood specimens from HAT patients and healthy endemic controls from D.R. Congo and Uganda. Both tests exhibited good sensitivity and specificity compared to the current diagnostic tests and may be valuable tools for sensitive and specific parasite detection in clinical specimens. These standardized molecular test formats open avenues for improved case detection, particularly in epidemiological studies and in disease diagnosis at reference centres. [ABSTRACT FROM AUTHOR]

Background: The development of arsenical and diamidine resistance in Trypanosoma brucei is associated with loss of drug uptake by the P2 purine transporter as a result of alterations in the corresponding T. brucei adenosine transporter 1 gene (TbAT1). Previously, specific TbAT1 mutant type alleles linked to melarsoprol treatment failure were significantly more prevalent in T. b. gambiense from relapse patients at Omugo health centre in Arua district. Relapse rates of up to 30% prompted a shift from melarsoprol to eflornithine (α-difluoromethylornithine, DFMO) as first-line treatment at this centre. The aim of this study was to determine the status of TbAT1 in recent isolates collected from T. b. gambiense sleeping sickness patients from Arua and Moyo districts in Northwestern Uganda after this shift in first-line drug choice. Methodology and results: Blood and cerebrospinal fluids of consenting patients were collected for DNA preparation and subsequent amplification. All of the 105 isolates from Omugo that we successfully analysed by PCR-RFLP possessed the TbAT1 wild type allele. In addition, PCR/RFLP analysis was performed for 74 samples from Moyo, where melarsoprol is still the first line drug; 61 samples displayed the wild genotype while six were mutant and seven had a mixed pattern of both mutant and wild-type TbAT1. The melarsoprol treatment failure rate at Moyo over the same period was nine out of 101 stage II cases that were followed up at least once. Five of the relapse cases harboured mutant TbAT1, one had the wild type, while no amplification was achieved from the remaining three samples. Conclusions/significance: The apparent disappearance of mutant alleles at Omugo may correlate with melarsoprol withdrawal as first-line treatment. Our results suggest that melarsoprol could successfully be reintroduced following a time lag subsequent to its replacement. A field-applicable test to predict melarsoprol treatment outcome and identify patients for whom the drug can still be beneficial is clearly required. This will facilitate cost-effective management of HAT in rural resource-poor settings, given that eflornithine has a much higher logistical requirement for its application. Author Summary: Human African trypanosomiasis (HAT) manifests as a chronic infection caused by Trypanosoma brucei gambiense, or as a more acute form due to T. b. rhodesiense. Both manifestations occur in Uganda and melarsoprol use against the former was jeopardised in the 1990s as reports of reduced efficacy increased to the point where it was dismissed as first-line treatment at some treatment centers. Previous work to elucidate possible mechanisms leading to melarsoprol resistance pointed to a P2 type adenosine transporter known to mediate melarsoprol uptake and previously shown to be mutated in significant numbers of patients not responding to the drug. Our present findings indicate that there is a low prevalence of mutants in foci where melarsoprol relapses are infrequent. In addition we observe that at the Omugo focus where the drug was withdrawn as first line over 6 years ago, the mutant alleles have disappeared, suggesting that drug pressure is responsible for fuelling their spread. Thus constant monitoring for mutants could play a key role in cost-effective HAT management by identifying which foci can still use the less logistically demanding melarsoprol as opposed to the alternative drug eflornithine. What is required now is a simple method for identifying such mutants at the point of care, enabling practitioners to make informed prescriptions at first diagnosis. [ABSTRACT FROM AUTHOR]

Objective To assess the application of allele-specific PCR (AS-PCR) as a fast, cheap and reliable method for detecting mutant TbAT1 associated with melarsoprol relapse in Trypanosoma brucei gambiense isolates from northwest Uganda. Methods A total of 105 trypanosome isolates were analysed using SfaN1 restriction fragment length polymorphism (RFLP) and AS-PCR, the former used as the gold standard. Sensitivity, specificity, positive and negative predictive values of AS-PCR as well as agreement between the tests were determined. Results Eleven trypanosome isolates had mutant TbAT1 while 94 exhibited the wild-type TbAT1 genes. There was a highly significant agreement between SfaN1 RFLP and AS-PCR with kappa and intra-class correlation values of 1.0. The sensitivity and specificity of AS-PCR were both 100%, while the positive and negative predictive values were found to be equal to 1.0. Cost and time analyses were performed and AS-PCR was 4.3 times cheaper than SfaN1 RFLP, in addition to the less time required for its execution. Conclusion AS-PCR should be the test of choice for screening for mutant TbAT1 in the ever-increasing numbers of field trypanosome isolates. [ABSTRACT FROM AUTHOR]

The human serum resistance-associated (SRA) gene was identified in 28 (80%) of the 35 T.b. rhodesiense trypanosomes from parasitologically confirmed sleeping sickness cases, using the primers designed by Radwanska and in 27 (77.1%) of the same 35 T.b. rhodesiense trypanosomes using the primers designed by Gibson. However, about 20% of the 35 T.b. rhodesiense trypanosomes could not be detected by SRA—polymerase chain reaction (PCR) even when an aliquot of the first PCR was used in the second PCR, indicating that the gene may be absent in those trypanosomes or the trypanosomes could be having another variant of SRA not detectable by these primers since three variants of SRA genes have so far been identified or the amount of trypanosomal DNA extracted from infected blood was too low to be detected. The trypanosome isolates that are SRA gene negative may indicate the presence of some T.b. rhodesiense trypanosomes with modified or lack SRA genes or simple loss of the SRA gene from the expression site in which it resides during antigenic variation. Analysis of trypanosomes derived from domestic animals showed that 79 (90.8%) of the 87 trypanosomes isolated from cattle were positive by Trypanosoma brucei (TBR)–PCR, indicating that they were Trypanozoon while 8 (9.2%) of the trypanosome isolates which were negative by TBR–PCR could be T. vivax, T. congolense, or T. theileri. When subjected to SRA–PCR, 10 (11.5%) of the 87 trypanosomes isolates derived from cattle were positive, indicating that there could be T.b. rhodesiense circulating in cattle, which is similar to the percentage of T.b. rhodesiense previously obtained in cattle in Serere, Soroti district. [ABSTRACT FROM AUTHOR]

African animal trypanosomiasis (AAT) is one of the major constraints to animal health and production in sub-Saharan Africa. To inform AAT control in Uganda and help advance along the progressive control pathway (PCP), we characterized AAT prevalence among eight host species in Uganda and explored factors that influence the prevalence variation between studies. We retrieved AAT prevalence publications (n = 2232) for Uganda (1980-2022) from five life sciences databases, focusing on studies specifying AAT detection methods, sample size, and the number of trypanosome-positive animals. Following PRISMA guidelines, we included 56 publications, and evaluated publication bias by the Luis Furuya-Kanamori (LFK) index. National AAT prevalence under DNA diagnostic methods for cattle, sheep and goats was 22.15%, 8.51% and 13.88%, respectively. Under DNA diagnostic methods, T. vivax was the most common Trypanosoma sp. in cattle (6.15%, 95% CI: 2.91-10.45) while T. brucei was most common among small ruminants (goats: 8.78%, 95% CI: 1.90-19.88, and sheep: 8.23%, 95% CI: 4.74-12.50, respectively). Northern and Eastern regions accounted for the highest AAT prevalence. Despite the limitations of this study (i.e., quality of reviewed studies, underrepresentation of districts/regions), we provide insights that could be used for better control of AAT in Uganda and identify knowledge gaps that need to be addressed to support the progressive control of AAT at country level and other regional endemic countries with similar AAT eco-epidemiology., (© 2023. The Author(s).)

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., Competing Interests: The authors have declared that no competing interests exist.

Competing Interests: Allan Mortensen and Georgina Bingham are employees of Vestergaard SA, which manufactures the Tiny Targets used in this project. They however play no role in implementation of the project.

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., (Copyright © 2020 The Authors. Published by Elsevier Inc. All rights reserved.)

Small ruminants are important to community livelihood in developing countries; however information on the role of hemoprotozoan parasites is scanty. The objective of the study was to determine hemoprotozoan parasitic prevalence in western Uganda and identify major areas associated with these infections. This was a cross sectional study conducted at the edge of Budongo Conservation Forest in Masindi district of western Uganda in which 712 small ruminants were sampled. Blood from the jugular vein was collected from caprines and ovines and placed in an EDTA tube, and transported to the laboratory for examination. Thin and thick smears were prepared and examined by microscopy for hemoprotozoan parasites, and DNA was extracted and examined by PCR for Trypanosoma spp. A total of 13 villages in Budongo sub-county were surveyed and the study showed that caprines were the major small ruminants of importance to the community. Prevalence of hemoprotozoan parasites was as follows; anaplasmosis (3.65%) > theileriosis (0.45%) > trypanosomiasis (0.15%) and babesiosis (0%) by microscopy. Infections were found in the young with the exception of Anaplasma spp. while coinfections of anaplasmosis and theileriosis were high. Molecular analysis showed an overall trypanosome prevalence of 9.27% (PCR), mainly due to Trypanosoma brucei and T. congolense forest. Villages with trypanosomiasis were found in lowlands and swamps. The current trypanosomiasis prevalence in small ruminants of Uganda was 10 times greater than that previously reported showing that the disease burden has increased overtime within Uganda. A prevalence of 0.14% (95% CI: 0.00, 0.78) for the SRA gene showed that small ruminants would be important reservoirs of infection to humans. Hemoprotozoan parasites are a threat to community livelihood in developing countries and the role of molecular diagnostic techniques in disease monitoring was re-emphasized by this study. Information on primary hosts involved in the propagation of hemoprotozoan parasites in Uganda would help streamline prospective disease surveillance and control efforts., (Copyright © 2019 Elsevier B.V. All rights reserved.)

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.

Background: The incidence of M. tuberculosis (MTB) and non tuberculous Mycobacterium species (NTMs) like M. avium and M. kansasii has increased due to Human Immunodeficiency Virus (HIV) epidemic. Therefore accurate, rapid and cost effective methods for the identification of these NTMs and MTB are greatly needed for appropriate TB management. Thus in this study we evaluated the performance of Lightcycler(®) Mycobacterium detection assay to detect MTB, M. avium and M. kansasii in sputum specimens., Methods: A total of 241 baseline minimally processed sputum specimens from individual adult TB suspected patients were analyzed by Mycobacterium detection assay (Real-time-PCR) on a LightCycler 480(®) while using liquid culture as a reference standard., Results: Real time PCR had a sensitivity of 100% (95% CI 96-100) and 100% (CI 19-100) for detection of MTB and M. avium respectively. Additionally the assay had a specificity of 99% (95% CI 96-99) and 95% (95% CI 91-97) for identification of MTB and M. avium respectively. The positive predictive value (PPV) for Real time PCR to identify MTB and M. avium among the specimens was 98% (95% CI 94-99) and 15% (95% CI 2-45) respectively. The kappa statistics for Real time PCR to identify MTB and M. avium was 0.9 (95% CI 0.9-1.0) and 0.3 (95% CI-0.03-0.5) respectively. The median time to detection for Real time PCR assay was 2 hours while overall median time to detection for MGIT-positive cultures was 8 days. The sample unit cost for Real time PCR was $ 12 compared to $ 20 for the reference liquid culture., Conclusion: The Light cycler(®) Mycobacterium detection assay rapidly and correctly identified MTB and M avium thus has the potential to be adopted in a clinical setting.

Confirmatory diagnosis of African trypanosomiasis relies on demonstration of parasites in body fluids by bright field microscopy. The parasitaemia in infected patients and animals is usually low, and concentration methods are used to try and increase the chances of seeing parasites. Recently, fluorescence microscopes using light-emitting diodes (LED) have been developed. Since they emit strong light, their use does not require a dark room, making field application a possibility. We have combined LED fluorescence microscopy with lysis of red blood cells (RBC) to improve the sensitivity and speed of detecting trypanosomes. In studies conducted at four centers in Uganda and the Democratic Republic of the Congo, parasitaemic blood was serially diluted and the RBCs lysed using commercial buffer. Samples were then concentrated by centrifugation, and different volumes of the sediment used to make thin and thick smears. Next, these were stained with acridine orange or Giemsa, and examined using an LED microscope under fluorescence or bright light, respectively. Detection of parasites was significantly improved by RBC lysis and concentration, regardless of the staining and microscopy method used. Further improvements were made when smears were prepared using larger volumes of sediment. The best results were obtained with thin smears prepared using 20 μl of sediment and stained with acridine orange. The time taken to see the first parasite was dramatically reduced when smears were examined by LED fluorescence microscopy, compared to bright light. LED fluorescence microscopy was found to be easier and requiring less visual effort than bright field microscopy. These studies demonstrate the potential for incremental improvement in detection of Trypanosoma brucei by combining LED fluorescence microscopy with RBC lysis and concentration. The lysis and concentration method may also be useful in sample preparation for other diagnostic tests for trypanosomiasis., (Copyright © 2011 Elsevier B.V. All rights reserved.)

We analyzed DNA eluted from FTA (Flinders Technology Associates) cards spotted with blood from human African trypanosomiasis (HAT) patients admitted at Lwala Hospital in eastern Uganda and Kaliua Health Centre in northwestern Tanzania. The aims were to evaluate loop-mediated isothermal amplification (LAMP) for detection of trypanosomal DNA in clinical samples and to characterize the infecting trypanosomes to the subspecies level. LAMP targeting the Trypanozoon conserved random inserted mobile element (RIME-LAMP) and that for the serum resistance-associated (SRA) gene (SRA-LAMP) were performed. For comparison, PCRs for the SRA gene specific for Trypanosoma brucei rhodesiense (SRA-PCR) and that to amplify the Trypanosoma brucei gambiense-specific surface glycoprotein (TgSGP-PCR) were done. Out of 128 samples analyzed, SRA-PCR was positive in 101 samples (78.9% sensitivity; 95% confidence interval [CI], 71.1 to 85.1%), SRA-LAMP was positive in 120 (93.8%; 95% CI, 88.2 to 96.8%), while RIME-LAMP revealed signals in 122 (95.3%; 95% CI, 90.2 to 97.8%). RIME-LAMP and SRA-LAMP were each significantly more sensitive than SRA-PCR (P values of 0.000 and 0.001, respectively; Fisher's exact test). There was poor agreement between RIME-LAMP and SRA-LAMP and the SRA-PCR, yielding kappa values of 0.31 and 0.40, respectively. Agreement between SRA-LAMP and RIME-LAMP was almost perfect (kappa value, 0.85; 95% CI, 0.64 to 1). All the 128 field samples were negative by TgSGP-PCR. Blood spots from three T. b. gambiense HAT cases from northwestern Uganda were positive by TgSGP-PCR and RIME-LAMP. PCR took five times longer to execute than LAMP. LAMP may be useful to monitor emerging HAT foci or to test travelers returning from countries where HAT is endemic. It should be evaluated in a case-control study to determine its utility as a HAT diagnostic.