East African Children Research Articles

Plasma metabolites in childhood Burkitt lymphoma cases and cancer-free controls in Uganda

IntroductionBurkitt lymphoma (BL) is an aggressive non-Hodgkin lymphoma associated with Plasmodium falciparum and Epstein-Barr virus, both of which affect metabolic pathways. The metabolomic patterns of BL is unknown.Materials and methodsWe measured 627 metabolites in pre-chemotherapy treatment plasma samples from 25 male children (6–11 years) with BL and 25 cancer-free area- and age-frequency-matched male controls from the Epidemiology of Burkitt Lymphoma in East African Children and Minors study in Uganda using liquid chromatography-tandem mass spectrometry. Unconditional, age-adjusted logistic regression analysis was used to estimate odds ratios (ORs) and their 95% confidence intervals (CIs) for the BL association with 1-standard deviation increase in the log-metabolite concentration, adjusting for multiple comparisons using false discovery rate (FDR) thresholds and Bonferroni correction.ResultsCompared to controls, levels for 42 metabolite concentrations differed in BL cases (FDR < 0.001), including triacylglyceride (18:0_38:6), alpha-aminobutyric acid (AABA), ceramide (d18:1/20:0), phosphatidylcholine ae C40:6 and phosphatidylcholine C38:6 as the top signals associated with BL (ORs = 6.9 to 14.7, P < 2.4✕10− 4). Two metabolites (triacylglyceride (18:0_38:6) and AABA) selected using stepwise logistic regression discriminated BL cases from controls with an area under the curve of 0.97 (95% CI: 0.94, 1.00).ConclusionOur findings warrant further examination of plasma metabolites as potential biomarkers for BL risk/diagnosis.

IFNL4 Genotypes and Risk of Childhood Burkitt Lymphoma in East Africa.

Interferon lambda 4 (IFN-λ4) is a novel type-III interferon that can be expressed only by carriers of the genetic variant rs368234815-dG within the first exon of the IFNL4 gene. Genetic inability to produce IFN-λ4 (in carriers of the rs368234815-TT/TT genotype) has been associated with improved clearance of hepatitis C virus (HCV) infection. The IFN-λ4-expressing rs368234815-dG allele (IFNL4-dG) is most common (up to 78%) in West sub-Saharan Africa (SSA), compared to 35% of Europeans and 5% of individuals from East Asia. The negative selection of IFNL4-dG outside Africa suggests that its retention in African populations could provide survival benefits, most likely in children. To explore this hypothesis, we conducted a comprehensive association analysis between IFNL4 genotypes and the risk of childhood Burkitt lymphoma (BL), a lethal infection-associated cancer most common in SSA. We used genetic, epidemiologic, and clinical data for 4,038 children from the Epidemiology of Burkitt Lymphoma in East African Children and Minors (EMBLEM) and the Malawi Infections and Childhood Cancer case-control studies. Generalized linear mixed models fit with the logit link controlling for age, sex, country, P. falciparum infection status, population stratification, and relatedness found no significant association between BL risk and 3 coding genetic variants within IFNL4 (rs368234815, rs117648444, and rs142981501) and their combinations. Because BL occurs in children 6-9 years of age who survived early childhood infections, our results suggest that additional studies should explore the associations of IFNL4-dG allele in younger children. This comprehensive study represents an important baseline in defining the health effects of IFN-λ4 in African populations.

Burkitt lymphoma risk shows geographic and temporal associations with Plasmodium falciparum infections in Uganda, Tanzania, and Kenya

Endemic Burkitt lymphoma (eBL) is a pediatric cancer coendemic with malaria in sub-Saharan Africa, suggesting an etiological link between them. However, previous cross-sectional studies of limited geographic areas have not found a convincing association. We used spatially detailed data from the Epidemiology of Burkitt Lymphoma in East African Children and Minors (EMBLEM) study to assess this relationship. EMBLEM is a case-control study of eBL from 2010 through 2016 in six regions of Kenya, Uganda, and Tanzania. To measure the intensity of exposure to the malaria parasite, Plasmodium falciparum, among children in these regions, we used high-resolution spatial data from the Malaria Atlas Project to estimate the annual number of P.falciparum infections from 2000 through 2016 for each of 49 districts within the study region. Cumulative P. falciparum exposure, calculated as the sum of annual infections by birth cohort, varied widely, with a median of 47 estimated infections per child by age 10, ranging from 4 to 315 infections. eBL incidence increased 39% for each 100 additional lifetime P. falciparum infections (95% CI: 6.10 to 81.04%) with the risk peaking among children aged 5 to 11 and declining thereafter. Alternative models using estimated annual P. falciparum infections 0 to 10 y before eBL onset were inconclusive, suggesting that eBL risk is a function of cumulative rather than recent cross-sectional exposure. Our findings provide population-level evidence that eBL is a phenotype related to heavy lifetime exposure to P. falciparum malaria and support emphasizing the link between malaria and eBL.

Human Leukocyte Antigen Contributes to Childhood Endemic Burkitt Lymphoma in Eastern Africa: A Case-Control Association Study

Background Endemic Burkitt lymphoma (eBL), which occurs at a disproportionately high rate among children in Africa, is associated with Plasmodium falciparum (Pf) malaria and Epstein-Barr virus (EBV). Genetic factors, including Human leukocyte antigen (HLA) system, are hypothesized to play a role in eBL by modulating the immune response or susceptibility to associated pathogens. However, the association between HLA and eBL has not been comprehensively evaluated to-date. Methods We investigated the association between HLA variants and eBL among 800 cases and 3,845 controls aged 0-15 years enrolled in two studies: the Epidemiology of Burkitt lymphoma in East African children and minors (EMBLEM) study conducted in Uganda, Tanzania and Kenya (2010-2016) and the Infections and Childhood Cancer Study conducted in Malawi (2005-2008). HLA alleles from eight classical HLA genes were imputed from genome wide data (Illumina 5 M array) with the SNP2HLA program using an updated multi-ancestry HLA reference panel (n=21,456). We demonstrated accurate imputation at 2-field resolution by comparing the imputed HLA alleles against HLA alleles obtained by sequence-based typing of 600 EMBLEM participants (200 cases and 400 controls) from Uganda. We performed country-specific association tests using generalized linear mixed models (GLMMs), adjusting for sex (genetic confirmed), age, falciparum positivity, and population structure using the top 3 principal components and accounting for genetic relatedness. Odds ratios (OR) were used to estimate the effect size for each HLA variant and significance was assessed by Wald tests. Meta-analyses were performed to obtain summary effects across countries. For 12 HLA alleles with previously reported associations with severe malaria (e.g., HLA-B*53), EBV antibodies (e.g., HLA-DRB1*15:01) or eBL (e.g., HLA-A*02), were hypothesized, apriori, that we would observe associations with eBL mirroring biologic effects of prior associations. Thus, a P value of 0.05 was considered sufficient for statistical significance. For all other associations, we adjusted for multiple testing with P<2.7×10-4 (correction for 187 tested HLA alleles) used to determine statistical significance for classical HLA alleles at 1- and 2-field resolution and P<1.1×10-6 (correction for 46,350 HLA variants) for all other variants in the HLA region. Conditional analyses were performed to identify HLA variants that independently influence eBL susceptibility. Results We found high concordance rates (>85%) of HLA imputation at all HLA class I and class II alleles of 2-field resolution. The frequency distribution of imputed HLA alleles was similar to that based on sequencing-based HLA typing in a subset of Ugandan participants (Pearson r=0.97). The expected associations between eBL and HLA alleles were only observed for HLA-A*02 and -B*41 based on a nominal p<0.05. More importantly, we found significant associations between eBL and HLA-DQA1*04:01 (adjusted odds ratio [aOR]= 1.61, 95% confidence interval [CI]=1.32 to 1.97, P=3.71×10-6) and SNP rs2040406, located in the HLA-DQA1 region (aOR=1.43 (95%CI=1.26 to 1.63), P=4.62×10-8) (Figure 1). No other HLA alleles or non-allele variants were significantly associated with eBL in conditional analysis on HLA-DQA*04:01 or rs2040406. The novel association with SNP rs2040406 corresponds to a single residue Gln53 within the HLA-DQA1 peptide-binding groove that tracks with rs2040406 (r2 = 0.88) and showed the strongest association among all residues (aOR=1.36, P=2.06×10-6). Conclusions We present a comprehensive investigation of the associations between eBL and HLA variants utilizing imputed HLA data. Consistent associations were observed for two eBL-based apriori-stipulated hypotheses (HLA-A*02 and-B*41); however, contrary to our expectations, the malaria-based apriori hypotheses were not confirmed. Our agnostic analysis identified new associations between eBL and variants in the HLA-DQB1/DQA1 region, thereby providing new directions to explore the underlying relationship between HLA, immunity and eBL risk. In particular, the association of eBL with rs2040406 was attributed to residue Gln53 within the HLA-DQA1 peptide-binding groove. These associations hint at functional effects, perhaps mediated by stronger binding of EBV glycoproteins. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Increased Frequency of Mosaic Chromosomal Alterations in Peripheral Blood Samples of Sub-Saharan African Children with Endemic Burkitt Lymphoma

Introduction: Burkitt lymphoma (BL) is an aggressive B-cell lymphoma with reciprocal IG∷MYC translocations and accounts for 50-75% of childhood cancers in sub-Saharan Africa. Mosaic chromosomal alterations (mCAs) are large, somatically acquired structural chromosomal alterations that are clonal and observed in detectable fractions of circulating mononuclear cells. mCAs have been linked to increased hematologic cancer risk, but their association with BL, a mononuclear B cell tumor, is unknown. We hypothesized that the frequency and distribution of mCAs in peripheral leukocytes would be elevated in children with BL, potentially serving as a biomarker of chromosomal instability relevant to BL risk. Methods: We investigated peripheral leukocyte DNA extracted from children enrolled in the Epidemiology of Burkitt Lymphoma in East African Children and Minors (EMBLEM) study (2010-2016) and the Infections and Childhood Cancer case-control study conducted in Malawi (2005-2010). We used genome-wide single nucleotide polymorphism (SNP) array data to investigate the frequency, type, and distribution of mCAs >2Mb in size detected in peripheral leukocytes of 931 pediatric BL cases with 3,645 healthy children in Uganda, Tanzania, and Kenya, and with 177 non-BL pediatric cancers in Malawi. Eagle2 and SHAPEIT4 software were used for phasing to infer haplotypes for SNP array data and WGS data. MoChA (v2014-05-14) was used to detect mCAs. Parallel analysis using similar methods was performed on whole genome sequences (WGS) from paired tumor-normal blood samples from two Ugandan BL patients from the BL Genome Sequencing Project (BLGSP) and from paired BL- and lymphoblastoid-derived cell lines. Results: Most samples (92%) were from the EMBLEM study, whereas 8% were from Malawi. The mean age of the BL cases and controls was similar (~7.4 years in both, P=0.60). Table 1 shows detected autosomal mCAs for BL cases and controls by country. Overall, a total of 250 autosomal mCAs were detected in 78 (8.4%) of the 931 BL cases. By comparison, a total of 188 autosomal mCAs were detected in 131 (3.4%) of the 3822 controls. Autosomal mCAs were detected in cell fractions ranging from 0.5% to 99.2% (Figure S1). The average number of autosomal mCAs per person (i.e., total of mCAs detected/number of individuals with mCAs detected) was higher in the BL cases compared to controls (3.21 [SD 3.24] versus 1.44 [SD 2.31], P-value=4.50×10-5. The higher frequency of mCAs in blood samples of children with BL compared to controls was independent of age, sex, P falciparum status, and country (OR=2.8, 95% CI=2.06-3.81, P-value=5.70×10-11, Table 1. mCAs on the X chromosome in girls were also associated with BL (5% versus 1.7%, OR=3.12, 95% CI=1.47-6.29, P-value=1.65×10-3). We did not observe an association for mCAs on the Y chromosome in boys. Analysis of paired tumor-normal samples confirmed the presence of the IG∷MYC translocation in tumor, but the same pipelines did not detect the translocation in the corresponding normal blood sample. In informative tumor-normal cases, some mCAs were only detected in tumor, others were detected in both tumor and normal blood, whereas others were detected mCAs in normal only suggesting some mCAs in blood could arise from pre-BL clones. mCAs only observed in BL cases were predominantly gains on chromosome 1q and 8 (Figure 1) which contain major drivers of Burkitt lymphomagenesis including PLEKHO1, MCL1, PSMB4, ILF2, HAX1, ATP8B2 and CKS1B.Conclusion: We report a high frequency of mCAs in healthy sub-Saharan African children and even higher frequency in pediatric BL cases. The mCAs in children with BL clustered on chromosomes 1q and 8 in regions that overlap with several significantly mutated BL genes, suggesting that mCAs may be potential biomarkers important in the natural history of BL. Using paired tumor-normal sample WGS data, we report both similar and unique mCAs in the paired samples suggesting pre-BL clones in blood. These methods are scalable to large cohorts with SNP array data to investigate health effects associated with mCAs in Africa. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

HBB rs334, ABO Rs8176703 and Plasmodium Falciparum Positivity at Enrollment Are Independently Associated with Lower Risk for Endemic Burkitt Lymphoma in Uganda, Tanzania, Kenya, and Malawi

Background Burkitt lymphoma (BL) is an aggressive B-cell lymphoma that accounts for 50-75% of childhood cancers in sub-Saharan Africa. Recurrent infection with Plasmodium (P.) falciparum is hypothesized to be causally linked to BL, but the epidemiological evidence is limited because it is based on exposures measured after BL onset. The prolonged period before BL onset makes traditional cohort studies impractical. To circumvent this problem, it was proposed that genetic variants that confer genetic resistance to P falciparum, such as heterozygotes for the HBB sickle allele (rs334), could be used to demonstrate a causal link with malaria. However, results from six studies have been conflicting, perhaps, reflecting effects of small sample size, design flaws in the studies or confounding. For example, co-inheritance with other genetic variants, such as a-thalassemia, have been reported to confound the epidemiological associations of severe malaria with rs334. Additionally, resistance against P. falciparum due to "premunition", defined as anti-clinical and anti-parasite resistance due to acquired immunity, could confound genetic associations with BL. To address the hypothesis of whether resistance to malaria protects against BL, we investigated the associations between BL and rs334 and 31 other single nucleotide polymorphisms (SNPs) plus α-thalassemia (hereafter, index malaria resistance SNPs/variants), and premunition, defined as P. falciparum positivity at enrollment, in cases enrolled in Uganda, Tanzania, Kenya, and Malawi, all designated as high malaria burden countries by the World Health Organization. Methods The study cohort consisted of 800 children (<16 years) with BL and 3845 children without BL enrolled in the Epidemiology of Burkitt Lymphoma in East African Children and Minors (EMBLEM) study (2010-2016) and the Infections and Childhood Cancer case-control study conducted in Malawi (2005-2010). P falciparum status was determined by thick-film microscopy, malaria antigen (HRP2/pLDH) rapid diagnostic test, or PCR at enrollment. Malaria protective SNPs were extracted from genome-wide data measured using Infinium Omni5Exome-4 v1.3 BeadChip (Illumina, San Diego, CA, USA). Genotyping for α3.7-thalassemia deletions (/-α3.7)utilized droplet digital polymerase chain reaction. We tested for associations between BL and the malaria index SNPs, a genotypes, and P. falciparum positivity using generalized linear mixed models, controlling for age, sex, and the first 3 population-specific principal components as fixed effects and for genetic relatedness (calculated from genome-wide data) as a random effect. We considered a p-value < 0.05 for statistical significance for malaria index SNPs. Results The mean age was similar for BL cases and controls (7.19 years versus 7.43 years, p= 0.07), but BL cases were more likely to be males (63.2% versus 52.1%, p£0.0001). P. falciparum positivity was lower in BL cases than controls (35.2% versus 48.3%, p<0.0001). Figure 1 shows the associations between 32 malaria index SNPs with BL and with P. falciparum positivity. Odds of BL were significantly decreased in presence of HBB rs334 (odds ratio [OR]= 0·687, 95% CI 0·533-0·885; p=0·0037) and ABO rs8176703 (0·591, 0·379-0·922; p=0.0203). Results for α-genotypes included the normal genotype (αα/αα), triplication (ααα/αα), -α3.7/αα, -α3.7/-α3.7deletions and unknown. Using αα/αα as the reference, the ORs for BL were 1.82, p=0.06 for ααα/αα; 1.07, p=0.469 for -α3.7/αα; and 0.78, p=0.15 for -α3.7/-α3.7. The odds of BL were significantly decreased with P. falciparum positivity in analyses without the two significant index SNPs (OR= 0.526, 95% CI 0.428-0.633, p=9.64x10-12) and changed minimally when the two SNPs were included (OR= 0·524, 0·435-0·630, p=7·291x10-12) . Conclusion Based on 4,581 children in four high malaria burden countries, we found reduced odds of BL among carriers of malaria index variants in HBB and ABO genes which suggests that these variants may protect against BL as well. Additionally, our findings of decreased odds of BL with P. falciparum positivity at enrollment suggest premunition may be protective against BL, independent of the significant associations with SNPs in HBB and ABO. Together, our findings further enhance the evidence of a causal role of malaria in BL; coordination between malaria and BL programs in countries with high malaria/BL burden should be strengthened. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Nodding syndrome: A key role for sources of nutrition?

Nodding Syndrome (NS) has occurred among severely food-stressed communities in northern Uganda and several other East African populations that, with their forced physical displacement, have resorted to nutritional support from available wild plants and fungi, some of which have neurotoxic potential. Among the latter is an agaric mushroom with an unknown content of hydrazine-generating agaritine, namely Agaricus bingensis, the unusually wide consumption of which may relate to the low serum levels of vitamin B6 in Ugandan NS subjects relative to controls. Hydrazine-related compounds induce patterns of DNA damage that promote neuropathological changes (tauopathy) reminiscent of those associated with established NS. While the cause of this childhood brain disease is unknown, we encourage increased attention to the role of malnutrition and B6 hypovitaminosis in the etiology of this devastating brain disease.

Pediatric speech-language pathology in East Africa: educational opportunities and availability of speech-language services

Objective This paper describes the current situation of the provision of speech-language services in East Africa. An overview of the educational opportunities, workforce density, communication rehabilitation planning, caseload, and ethical and/or cultural challenges is provided. Method A narrative review was performed. All papers referring to communication disorders in East African countries were identified from EMBASE, PubMed, ISI Web of Science: Science Citation Index Expanded, and ISI Web of Science: Conference Proceedings Citation Index-Science using the following search terms: speech therapy, speech pathology, speech-language pathology, speech-language rehabilitation, speech-language therapy, logopedics, East Africa, communication disorders, communication difficulties. Two concepts were combined with a Boolean operator ‘AND’. Results At present, there is an alarming shortage of speech-language pathologists in East Africa. This issue is mainly related to the limited availability of educational programs in speech-language pathology. Although more and more universities are beginning to offer such educational programs, the number of available speech-language pathologists is still too low to meet the needs of all children with communication disorders. In addition, speech-language pathologists are often faced with different cultural and ethical obstacles when they provide intervention to East African children with communication disorders. These speech-language pathologists are therefore challenged to take a broader view of their role as a therapist. These alternative roles often include training of other individuals in the context of community-based rehabilitation. Conclusion In the future, it will be important to carefully consider the development of sustainable learning opportunities in East African contexts. Initiatives, for example, “train-the-trainer” workshops, are needed to support the speechlanguage pathologists. Existing models like the communication disability model can aid to create holistic frameworks for the development of sustainable speech-language services in the East African countries.

Epstein-Barr Virus in Burkitt Lymphoma in Africa Reveals a Limited Set of Whole Genome and LMP-1 Sequence Patterns: Analysis of Archival Datasets and Field Samples From Uganda, Tanzania, and Kenya.

Epstein-Barr virus (EBV) is associated with endemic Burkitt lymphoma (eBL), but the contribution of EBV variants is ill-defined. Studies of EBV whole genome sequences (WGS) have identified phylogroups that appear to be distinct for Asian versus non-Asian EBV, but samples from BL or Africa, where EBV was first discovered, are under-represented. We conducted a phylogenetic analysis of EBV WGS and LMP-1 sequences obtained primarily from BL patients in Africa and representative non-African EBV from other conditions or regions using data from GenBank, Sequence Read Archive, or Genomic Data Commons for the Burkitt Lymphoma Genome Sequencing Project (BLGSP) to generate data to support the use of a simpler biomarker of geographic or phenotypic associations. We also investigated LMP-1 patterns in 414 eBL cases and 414 geographically matched controls in the Epidemiology of Burkitt Lymphoma in East African children and minors (EMBLEM) study using LMP-1 PCR and Sanger sequencing. Phylogenetic analysis revealed distinct genetic patterns of African versus Asian EBV sequences. We identified 281 single nucleotide variations (SNVs) in LMP-1 promoter and coding region, which formed 12 unique patterns (A to L). Nine patterns (A, AB, C, D, F, I, J, K and L) predominated in African EBV, of which four were found in 92% of BL samples (A, AB, D, and H). Predominant patterns were B and G in Asia and H in Europe. EBV positivity in peripheral blood was detected in 95.6% of EMBLEM eBL cases versus 79.2% of the healthy controls (odds ratio [OR] =3.83; 95% confidence interval 2.06-7.14). LMP-1 was successfully sequenced in 66.7% of the EBV DNA positive cases but in 29.6% of the controls (ORs ranging 5-11 for different patterns). Four LMP-1 patterns (A, AB, D, and K) were detected in 63.1% of the cases versus 27.1% controls (ORs ranges: 5.58-11.4). Dual strain EBV infections were identified in WGS and PCR-Sanger data. In conclusion, EBV from Africa is phylogenetically separate from EBV in Asia. Genetic diversity in LMP-1 formed 12 patterns, which showed promising geographic and phenotypic associations. Presence of multiple strain infection should be considered in efforts to refine or improve EBV markers of ancestry or phenotype.Lay SummaryEpstein-Barr virus (EBV) infection, a ubiquitous infection, contributes to the etiology of both Burkitt Lymphoma (BL) and nasopharyngeal carcinoma, yet their global distributions vary geographically with no overlap. Genomic variation in EBV is suspected to play a role in the geographical patterns of these EBV-associated cancers, but relatively few EBV samples from BL have been comprehensively studied. We sought to compare phylogenetic patterns of EBV genomes obtained from BL samples in Africa and from tumor and non-tumor samples from elsewhere. We concluded that EBV obtained from BL in Africa is genetically separate from EBV in Asia. Through comprehensive analysis of nucleotide variations in EBV’s LMP-1 gene, we describe 12 LMP-1 patterns, two of which (B and G) were found mostly in Asia. Four LMP-1 patterns (A, AB, D, and F) accounted for 92% of EBVs sequenced from BL in Africa. Our results identified extensive diversity of EBV, but BL in Africa was associated with a limited number of variants identified, which were different from those identified in Asia. Further research is needed to optimize the use of PCR and sequencing to study LMP-1 diversity for classification of EBV variants and for use in epidemiologic studies to characterize geographic and/or phenotypic associations of EBV variants with EBV-associated malignancies, including eBL.

Plasma EBV DNA: A Promising Diagnostic Marker for Endemic Burkitt Lymphoma.

Endemic Burkitt lymphoma (eBL) is the most common childhood cancer in regions of equatorial Africa where P. falciparum malaria is holoendemic. The tumor is consistently associated with Epstein-Barr virus (EBV). Screening for EBV DNA in plasma in a high-risk population in Hong Kong has been shown to be useful in facilitating the early diagnosis of nasopharyngeal carcinoma, another EBV-associated tumor. Here, we investigate plasma EBV as a diagnostic marker for eBL in children in Uganda. We studied plasma specimens from 25 children with eBL and 25 controls matched for age (<3-16 years), gender and geography, including many with asymptomatic P. falciparum infection. These specimens were previously collected under the auspices of the EMBLEM (Epidemiology of Burkitt lymphoma in East African children and minors) study. After cell-free DNA isolation, plasma EBV DNA was measured using a quantitative PCR assay that amplifies the large internal repeats of the EBV genome. All children with eBL had measurable plasma EBV, as compared to 84% of control children. The median plasma EBV DNA level was 5.23 log10 copies/mL (interquartile range 3.54-6.08 log10 copies/mL) in children with eBL. In contrast, the median plasma EBV DNA level was 0.37 log10 copies/mL (interquartile range 0.18-1.05 log10 copies/mL) in children without lymphoma. An EBV threshold of 2.52 log10 copies/mL yielded a sensitivity of.88 and a specificity of 1. The estimated AUC was 0.936 (95% CI: 0.8496 – 1.00) for the corresponding ROC curve. Plasma EBV copy number did not depend on age, gender, or malaria screening status. However, two control children with asymptomatic P. falciparum infection and parasitemia also had high plasma EBV copy number. Our analysis suggests that measurements of EBV copy number in plasma may be useful in identifying children with eBL versus control children. A promising area for future research is the differentiation of high copy number associated with tumor versus high copy number associated with asymptomatic parasitemia.

Prevalence of rheumatic heart disease among school children in East Africa: a systematic review and meta-analysis.

Introductionconducting researches and estimating the prevalence of rheumatic heart disease in school children is crucial to develop school-based strategies targeted to decrease the prevalence of this disease. Therefore, this systematic review and meta-analysis were aimed to estimate the overall prevalence of rheumatic heart disease (RHD) among school children in East Africa.MethodsPubMed/MEDLINE, Google Scholar, Cochrane review, African Journals Online and African Index Medicus databases were searched to identify relevant research articles. The overall prevalence of rheumatic heart disease was pooled based on the weighted inverse variance random-effects model at a 95% confidence interval. The presence of heterogeneity, sensitivity analysis and presence of publication bias was tested. Results were presented with narrative synthesis, tables and forest plots.Resultsa total of thirteen research articles were included in the final analysis. The pooled prevalence of rheumatic heart disease in East African school children was 1.79% (17.9 cases per 1000 children (95% CI=11.6, 24.2; I2=95.1%; p<0.001)). From the subgroup analysis conducted by publication year, a lower prevalence of RHD in school children was reported among studies published after 2015 (six studies; overall prevalence=1.17% (11.7 cases per 1000 school children); with 95% CI=0.60, 1.73%; I2=88.8%; p<0.001). Additionally, the horn of Africa was found to have the lowest prevalence of RHD in school children among East African countries (six studies; overall prevalence=1.59% (15.9 cases per 1000 school children); with 95% CI=0.68, 2.51%; I2=94.2%; p<0.001).Conclusionthe prevalence of rheumatic heart disease (RHD) among school children in East Africa was considerably higher than the results from high-income countries. Therefore, community education on strep throat and its complications should be implemented through mass media. Rheumatic heart disease preventive strategies should be integrated with schools to reduce the prevalence of RHD among school children.

Endemic Burkitt lymphoma: a complication of asymptomatic malaria in sub-Saharan Africa based on published literature and primary data from Uganda, Tanzania, and Kenya

BackgroundEndemic Burkitt lymphoma (eBL) is an aggressive B cell non-Hodgkin lymphoma associated with antigenic stimulation from Plasmodium falciparum malaria. Whether eBL risk is related to malaria parasite density is unknown. To address this issue, children with eBL, asymptomatic and clinical malaria, as a surrogate of malaria parasite density, were assessed.MethodsMalaria-related laboratory results (parasite density, haemoglobin, platelet count, and white cell count [WBC]) count) were compiled for 4019 eBL cases and 80,532 subjects evaluated for asymptomatic malaria or clinical malaria (severe malaria anaemia, hyperparasitaemia, cerebral malaria, malaria prostration, moderate malaria, and mild malaria) in 21 representative studies published in Africa (mostly East Africa) and 850 eBL cases and 2878 controls with primary data from the Epidemiology of Burkitt Lymphoma in East African Children and Minors (EMBLEM) case–control study in Uganda, Tanzania, and Kenya. The average values of malaria-related laboratory results were computed by condition and trends across single-year age groups were assessed using regression and spline models.ResultsOverall, malaria infection or malaria was diagnosed in 37,089 of children compiled from the literature. Children with eBL and asymptomatic parasitaemia/antigenaemia, but not those with clinical malaria, were closest in their mean age (age 7.1–7.2 vs. 7.4–9.8 years), haemoglobin level (10.0–10.4 vs. 11.7–12.3 g/dL), malaria parasite density (2800 vs. 1827–7780 parasites/µL), platelet count (347,000–353,000 vs. 244,000–306,000 platelets/µL), and WBC count (8180–8890 vs. 7100–7410 cells/µL). Parasite density in these two groups peaked between four to five years, then decreased steadily thereafter; conversely, haemoglobin showed a corresponding increase with age. Children with clinical malaria were markedly different: all had an average age below 5 years, had dramatically elevated parasite density (13,905–869,000 parasites/µL) and dramatically decreased platelet count (< 159,000 platelets/µL) and haemoglobin (< 7 g/dL).ConclusionseBL and asymptomatic parasitaemia/antigenaemia, but not clinical malaria, were the most similar conditions with respect to mean age and malaria-related laboratory results. These results suggest that children with asymptomatic parasitaemia/antigenaemia may be the population at risk of eBL.

Continuous glucose monitoring assessment of metabolic control in east African children and young adults with type 1 diabetes: A pilot and feasibility study.

BackgroundFor individuals with type 1 diabetes (T1D) in East Africa and other low‐income regions, the last decade has seen substantial gains in access to insulin and trained healthcare providers, yet metabolic control remains poor.MethodsThe objective was to determine the feasibility of continuous glucose monitoring (CGM) and to gather baseline metabolic data for future power analysis in Ugandan and Kenyan youth with T1D using a Freestyle Libre Pro blinded CGM.ResultsOf 78 participants recruited, four sensors fell off and six patients did not return, leaving 68 evaluable subjects. Average age was 16 ± 5 (range 4‐26) years, 43% female. Average diabetes duration was 7 ± 5 years, insulin dose 0.9 ± 0.3 U/kg/d, and number of fingerstick glucose levels per day 2.1 ± 1.1. All were on human insulin. Point‐of‐care HbA1c was 10.9 ± 2.7% (96 ± 30 mmol/mol). Mean number of sensor days was 13 ± 3; >90% wore the sensor for ≥10 days. Mean glucose was 231 ± 86 mg/dL (12.8 ± 4.8 mmol/L). Only 30 ± 19% of time was spent in the target range (70‐180 mg/dL; 3.9‐10 mmol/L), and 7 ± 8% of time was spent in hypoglycaemia (glucose <55 mg/dL, 3.0 mmol/L). Hypoglycaemia occurred in 81% of participants, averaging five events/wk with an average duration of 140 ± 79 minutes/event.ConclusionsDespite significant diabetes care improvements, East African youth with T1D have poor metabolic control with chronic hyper‐ and hypoglycaemia, placing them at high risk for serious acute and chronic complications. This study demonstrates the feasibility of CGM use in this population and provides baseline metabolic data that will be used to inform a future intervention study.

SAT-246 CHRONIC KIDNEY DISEASE IMPACTS HEALTH-RELATED QUALITY OF LIFE OF CHILDREN IN UGANDA, EAST AFRICA

Limited data exists about the epidemiology of kidney disease in sub-Saharan East African children or impact of chronic kidney disease (CKD) on health-related quality of life (HRQoL). Our objective was to describe types of renal disease, compare HRQoL in Ugandan children with CKD and children with benign or resolving renal disease (non-CKD), and assess for predictors of HRQoL. This cross-sectional study of children aged 0-18 years assessed demographic, socioeconomic and clinical data and surveyed HRQoL using the Pediatric Quality of Life Core ScaleTM (PedsQL) (scoring scale 0-100) for 4 domains (physical, emotional, social and school) and overall HRQoL. CKD and non-CKD participant scores were compared using unpaired t-test. Predictors of HRQoL were evaluated using linear and logistic regression analyses. A total of 149 children were enrolled (71 in CKD group; 78 in non-CKD group; median age 9 years; male 63%). Primary renal diseases included nephrotic syndrome 83 (56%), glomerulonephritis 25 (17%), obstructive uropathy 22 (15%), cystic kidney disease 6 (3.7%), acute kidney injury 2 (1.3%), hypertension 2 (1.3%), human immunodeficiency virus-associated nephropathy 1 (0.7%), and unknown renal disease 8 (5%). Congenital anomalies of the kidney and urinary tract (CAKUT) accounted for 39% (28/71) of CKD etiologies. In the CKD group, 46% were at stage 5 with 10%, 14%, 16%, and 14% at stages 4, 3, 2 and 1, respectively. Among school-eligible children, 34% (19/56) with CKD did not attend school whereas all non-CKD did (p<0.0001); CKD children enrolled in school missed on average 14 days per month. CKD children had significantly lower HRQoL domain scores. Mean overall HRQoL score by child report was 57 for CKD vs. 86 for non-CKD (p<0.0001), similar to caregiver report 63 for CKD vs. 86 for non-CKD (p<0.0001). Predictors of lower HRQoL among CKD children were advanced CKD stages, primary caregiver other than child’s parent, low hemoglobin, and vitamin D deficiency. As in other parts of the world, CAKUT was the main cause of CKD in Ugandan children. The majority of children presented at late CKD stages. HRQoL in CKD was much lower than in non-CKD children. Only two thirds of school-eligible CKD children actually attended school. Low hemoglobin and vitamin D deficiency were potentially modifiable factors associated with low HRQoL. Simple interventions with iron, erythropoietin stimulating agent, and vitamin D supplementation might lead to improved HRQoL.

Risk factors for Burkitt lymphoma in East African children and minors: A case-control study in malaria-endemic regions in Uganda, Tanzania and Kenya.

Endemic Burkitt lymphoma (eBL) is the most common childhood cancer in sub-Saharan African countries, however, few epidemiologic studies have been undertaken and none attempted enrolling cases from multiple countries. We therefore conducted a population-based case-control study of eBL in children aged 0-15 years old in six regions in Northern Uganda, Northern Tanzania and Western Kenya, enrolling 862 suspected cases and 2,934 population controls (response rates 98.5-100%), and processing ~40,000 vials of samples using standardized protocols. Risk factor questionnaires were administered, and malaria period prevalence was measured using rapid diagnostic tests (RDTs). A total of 80.9% of the recruited cases were diagnosed as eBL; 61.4% confirmed by histology. Associations with eBL risk were computed using logistic regression models adjusted for relevant confounders. Associations common in at least two countries were emphasized. eBL risk was decreased with higher maternal income and paternal education and elevated with history of inpatient malaria treatment >12 months before enrollment. Reporting malaria-attributed fever up to 6 months before enrollment and malaria-RDT positivity at enrollment were associated with decreased eBL risk. Conversely, reporting exposure to mass malaria suppression programs (e.g., indoor residual insecticide) was associated with elevated risk. HIV seropositivity was associated with elevated eBL risk, but the relative impact was small. The study shows that it is feasible to conduct networked, multisite population-based studies of eBL in Africa. eBL was inversely associated with socioeconomic status, positively associated with inpatient malaria treatment 12 months ago and with living in areas targeted for malaria suppression, which support a role of malaria in eBL.

Enteric virome of Ethiopian children participating in a clean water intervention trial.

BackgroundThe enteric viruses shed by different populations can be influenced by multiple factors including access to clean drinking water. We describe here the eukaryotic viral genomes in the feces of Ethiopian children participating in a clean water intervention trial.Methodology/principal findingsFecal samples from 269 children with a mean age of 2.7 years were collected from 14 villages in the Amhara region of Ethiopia, half of which received a new hand-dug water well. Feces from these villages were then analyzed in 29 sample pools using viral metagenomics. A total of 127 different viruses belonging to 3 RNA and 3 DNA viral families were detected. Picornaviridae family sequence reads were the most commonly found, originating from 14 enterovirus and 6 parechovirus genotypes plus multiple members of four other picornavirus genera (cosaviruses, saliviruses, kobuviruses, and hepatoviruses). Picornaviruses with nearly identical capsid VP1 were detected in different pools reflecting recent spread of these viral strains. Next in read frequencies and positive pools were sequences from the Caliciviridae family including noroviruses GI and GII and sapoviruses. DNA viruses from multiple genera of the Parvoviridae family were detected (bocaviruses 1–4, bufavirus 3, and dependoparvoviruses), together with four species of adenoviruses and common anelloviruses shedding. RNA in the order Picornavirales and CRESS-DNA viral genomes, possibly originating from intestinal parasites or dietary sources, were also characterized. No significant difference was observed between the number of mammalian viruses shed from children from villages with and without a new water well.ConclusionsWe describe an approach to estimate the efficacy of potentially virus transmission-reducing interventions and the first complete (DNA and RNA viruses) description of the enteric viromes of East African children. A wide diversity of human enteric viruses was found in both intervention and control groups. Mammalian enteric virome diversity was not reduced in children from villages with a new water well. This population-based sampling also provides a baseline of the enteric viruses present in Northern Ethiopia against which to compare future viromes.

Piloting a Developmental Screening Tool Adapted for East African Children.

There is a need for developmental screening that is easily administered in resource-poor settings. We hypothesized that known risk factors would predict failed developmental screening on an adapted screening tool in East African children living in poverty. The sample included 100 healthy Ugandan children aged 6–59 months. We adapted a parent-reported developmental screener based on the Child Development Review chart. The primary outcome was failure to meet age-appropriate milestones for any developmental domain. Venous blood was analyzed for lead, and caregivers completed a demographics questionnaire. We used multivariate logistic regression models to determine if elevated blood lead and stunting predicted failure on the screener, controlling for maternal education level, age in months past the lower bound of the child’s developmental age group, and absence of home electricity. In the sample, 14% (n = 14) of children failed one or more milestones on the screener. Lead levels or stunting did not predict failing the screener after controlling for covariates. Though this tool was feasibly administered, it did not demonstrate preliminary construct validity and is not yet recommended for screening in high-risk populations. Future research should include a larger sample size and cognitive interviews to ensure it is contextually relevant.

Coinfection with Enteric Pathogens in East African Children with Acute Gastroenteritis-Associations and Interpretations.

Enteric coinfections among children in low-income countries are very common, but it is not well known if specific pathogen combinations are associated or have clinical importance. In this analysis, feces samples from children in Rwanda and Zanzibar less than 5 years of age, with (N = 994) or without (N = 324) acute diarrhea, were analyzed by real-time polymerase chain reaction targeting a wide range of pathogens. Associations were investigated by comparing co-detection and mono-detection frequencies for all pairwise pathogen combinations. More than one pathogen was detected in 840 samples (65%). A negative association (coinfections being less common than expected from probability) was observed for rotavirus in combination with Shigella, Campylobacter, or norovirus genogroup II, but only in patients, which is statistically expected for agents that independently cause diarrhea. A positive correlation was observed, in both patients and controls, between Ct (threshold cycle) values for certain virulence factor genes in enteropathogenic Escherichia coli (EPEC) (eae and bfpA) and toxin genes in enterotoxigenic E. coli (eltB and estA), allowing estimation of how often these genes were present in the same bacteria. A significant positive association in patients only was observed for Shigella and EPEC-eae, suggesting that this coinfection might interact in a manner that enhances symptoms. Although interaction between pathogens that affect symptoms is rare, this work emphasizes the importance and difference in interpretation of coinfections depending on whether they are positively or negatively associated.

High rates of viral suppression in adults and children with high CD4+ counts using a streamlined ART delivery model in the SEARCH trial in rural Uganda and Kenya.

Introduction: The 2015 WHO recommendation of antiretroviral therapy (ART) for all HIV-positive persons calls for treatment initiation in millions of persons newly eligible with high CD4+ counts. Efficient and effective care models are urgently needed for this population. We evaluated clinical outcomes of asymptomatic HIV-positive adults and children starting ART with high CD4+ counts using a novel streamlined care model in rural Uganda and Kenya.Methods: In the 16 intervention communities of the HIV test-and-treat Sustainable East Africa Research for Community Health Study (NCT01864603), all HIV-positive individuals irrespective of CD4 were offered ART (efavirenz [EFV]/tenofovir disoproxil fumarate + emtricitabine (FTC) or lamivudine (3TC). We studied adults (≥fifteen years) with CD4 ≥ 350/μL and children (two to fourteen years) with CD4 > 500/μL otherwise ineligible for ART by country guidelines. Clinics implemented a patient-centred streamlined care model designed to reduce patient-level barriers and maximize health system efficiency. It included (1) nurse-conducted visits with physician referral of complex cases, (2) multi-disease chronic care (including for hypertension/diabetes), (3) patient-centred, friendly staff, (4) viral load (VL) testing and counselling, (5) three-month return visits and ART refills, (6) appointment reminders, (7) tiered tracking for missed appointments, (8) flexible clinic hours (outside routine schedule) and (9) telephone access to clinicians. Primary outcomes were 48-week retention in care, viral suppression (% with measured week 48 VL ≤ 500 copies/mL) and adverse events.Results: Overall, 972 HIV-positive adults with CD4+ ≥ 350/μL initiated ART with streamlined care. Patients were 66% female and had median age thirty-four years (IQR, 28–42), CD4+ 608/μL (IQR, 487–788/μL) and VL 6775 copies/mL (IQR, <500–37,003 c/mL). At week 48, retention was 92% (897/972; 2 died/40 moved/8 withdrew/4 transferred care/21/964 [2%] were lost to follow-up). Viral suppression occurred in 778/838 (93%) and 800/972 (82%) in intention-to-treat analysis. Grade III/IV clinical/laboratory adverse events were rare: 95 occurred in 74/972 patients (7.6%). Only 8/972 adults (0.8%) switched ART from EFV to lopinavir (LPV) (n = 2 for dizziness, n = 2 for gynaecomastia, n = 4 for other reasons). Among 83 children, week 48 retention was 89% (74/83), viral suppression was 92% (65/71) and grade III/IV adverse events occurred in 4/83 (4.8%).Conclusions: Using a streamlined care model, viral suppression, retention and ART safety were high among asymptomatic East African adults and children with high CD4+ counts initiating treatment.Clinical Trial Number: NCT01864603

Molecular markers of anti-malarial drug resistance in Central, West and East African children with severe malaria

BackgroundThe Plasmodium falciparum multidrug resistance 1 (PfMDR1), P. falciparum Ca2+-ATPase (PfATP6) and Kelch-13 propeller domain (PfK13) loci are molecular markers of parasite susceptibility to anti-malarial drugs. Their frequency distributions were determined in the isolates collected from children with severe malaria originating from three African countries.MethodsSamples from 287 children with severe malaria [(Gabon: n = 114); (Ghana: n = 89); (Kenya: n = 84)] were genotyped for pfmdr1, pfatp6 and pfk13 loci by DNA sequencing and assessing pfmdr1 copy number variation (CNV) by real-time PCR.ResultsPfmdr1-N86Y mutation was detected in 48, 10 and 10% in Lambaréné, Kumasi and Kisumu, respectively. At codon 184, the prevalence of the mutation was 73% in Lambaréné, 63% in Kumasi and 49% Kisumu. The S1034C and N1042D variants were absent at all three sites, while the frequency of the D1246Y mutation was 1, 3 and 13% in Lambaréné, Kumasi and Kisumu, respectively. Isolates with two pfmdr1 gene copy number predominantly harboured the N86Y wild-type allele and were mostly found in Kumasi (10%) (P < 0.0001). Among the main pfmdr1 haplotypes (NFD, NYD and YFD), NYD was associated with highest parasitaemia (P = 0.04). At the pfatp6 locus, H243Y and A623E mutations were observed at very low frequency at all three sites. The prevalence of the pfatp6 E431K variant was 6, 18 and 17% in Lambaréné, Kumasi and Kisumu, respectively. The L263E and S769N mutations were absent in all isolates. The pfk13 variants associated with artemisinin resistance in Southeast Asia were not observed. Eleven novel substitutions in the pfk13 locus occurring at low frequency were observed.ConclusionsArtemisinins are still highly efficacious in large malaria-endemic regions though declining efficacy has occurred in Southeast Asia. The return of chloroquine-sensitive strains following the removal of drug pressure is observed. However, selection of wild-type alleles in the multidrug-resistance gene and the increased gene copy number is associated with reduced lumefantrine sensitivity. This study indicates a need to constantly monitor drug resistance to artemisinin in field isolates from malaria-endemic countries.