Longitudinal wastewater-based surveillance of SARS-CoV-2 during 2023 in Ethiopia.

Introduction: Although wastewater-based epidemiology (WBE) successfully functioned as a tool for monitoring the coronavirus disease 2019 (COVID-19) pandemic globally, relatively little is known about its utility in low-income countries. This study aimed to quantify severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA in wastewater, estimate the number of infected individuals in the catchment areas, and correlate the results with the clinically reported COVID-19 cases in Addis Ababa, Ethiopia. Methods: A total of 323 influent and 33 effluent wastewater samples were collected from three Wastewater Treatment Plants (WWTPs) using a 24-h composite Moore swab sampling method from February to November 2023. The virus was captured using Ceres Nanotrap® Enhancement Reagent 2 and Nanotrap® Microbiome A Particles, and then nucleic acids were extracted using the Qiagen QIAamp Viral RNA Mini Kit. The ThermoFisher TaqPath™ COVID-19 kit was applied to perform real-time reverse transcriptase polymerase chain reaction (qRT-PCR) to quantify the SARS-CoV-2 RNA. Wastewater viral concentrations were normalized using flow rate and number of people served. In the sampling period, spearman correlation was used to compare the SARS-CoV-2 target gene concentration to the reported COVID-19 cases. The numbers of infected individuals under each treatment plant were calculated considering the target genes' concentration, the flow rate of treatment plants, a gram of feces per person-day, and RNA copies per gram of feces. Results: SARS-CoV-2 was detected in 94% of untreated wastewater samples. All effluent wastewater samples (n = 22) from the upflow anaerobic sludge blanket (UASB) reactor and membrane bioreactor (MBR) technology were SARS-COV-2 RNA negative. In contrast, two out of 11 effluents from Waste Stabilization Pond were found positive. Positive correlations were observed between the weekly average SARS-CoV-2 concentration and the cumulative weekly reported COVID-19 cases in Addis Ababa. The estimated number of infected people in the Kality Treatment catchment area was 330 times the number of COVID-19 cases reported during the study period in Addis Ababa. Discussion: This study revealed that SARS-CoV-2 was circulating in the community and confirmed previous reports of more asymptomatic COVID-19 cases in Ethiopia. Additionally, this study provides further evidence of the importance of wastewater-based surveillance in general to monitor infectious diseases in low-income settings. Conclusion: Wastewater-based surveillance of SARS-CoV-2 can be a useful method for tracking the increment of COVID-19 cases before it spreads widely throughout the community.

Diversity of culturable bacterial isolates and their potential as antimicrobial against human pathogens from Afar region, Ethiopia.

Antimicrobial resistance is a growing global concern exacerbated by the scarcity of new medications and resistance to current antibiotics. Microbes from unexplored habitats are promising sources of natural products to combat this challenge. This study aimed to isolate bacteria producing secondary metabolites and assess their antimicrobial efficacy against human pathogens. Soil and liquid samples were collected from Afar region, Ethiopia. Bacterial isolates were obtained using standard serial dilution techniques. Antimicrobial activity was evaluated using agar plug and well diffusion methods. matrix-assisted laser desorption/ionization time-of-flight-mass spectrometry (MALDI-TOF MS) and whole-genome sequencing (WGS) were conducted for the isolate exhibiting the highest antimicrobial activity. Secondary metabolites were extracted and analyzed using gas chromatography-mass spectra (GC-MS). In this study, 301 bacteria isolates were identified, of which 68 (22.6%) demonstrated antagonistic activity against at least one reference pathogen. Whole-genome sequencing revealed that Sl00103 belongs to the genus Bacillus, designated as Bacillus sp. Sl00103. The extract of Sl00103 showed zones of inhibition ranging between 17.17 ± 0.43 and 26.2 ± 0.4 mm against bacterial pathogens and 19.5 ± 0.44 to 21.0 ± 1.01 mm against Candida albicans. GC-MS analysis of ethyl acetate and n-hexane extracts identified major compounds including (R,R)-butane-2,3-diol; 3-isobutylhexahydropyrrolo[1,2a] pyrazine-1,4-dione; cyclo(L-prolyl-L-valine); and tetradecanoic acid, 12-methyl-, methyl ester; hexadecanoic acid, methyl ester among other. In conclusion, this study isolated several promising bacterial strains from the Afar region in Ethiopia, with strain Sl00103 (Bacillus sp. Sl00103) demonstrating notable antimicrobial and antioxidant activities and warranting further studies. IMPORTANCE: Antimicrobial resistance (AMR) is an escalating global health threat affecting humans, animals, and the environment, underscoring the urgent need for alternative pathogen control methods. Natural products, particularly secondary metabolites from bacteria, continue to be a vital source of antibiotics. However, microbial habitats and metabolites in Africa remain largely unexplored. In this study, we isolated and screened bacteria from Ethiopia's Afar region, characterized by extreme conditions like high temperatures, volcanic activity, high salinity, and hot springs to identify potential bioactive compounds. We discovered diverse bacterial isolates with antimicrobial activity against various pathogens, including strain Sl00103 (Bacillus sp. Sl00103), which demonstrated significant antimicrobial and antioxidant activities. GC-MS analysis identified several antimicrobial compounds, highlighting strain Sl00103 as a promising source of secondary metabolites with potential pharmaceutical applications and warranting further investigation.

Genomic characterization of Dengue virus circulation in Ethiopia.

In Ethiopia, dengue virus (DENV) infections have been reported in several regions, however, little is known about the circulating genetic diversity. Here, we conducted clinical surveillance for DENV during the 2023 nationwide outbreak and sequenced DENV whole genomes for the first time in Ethiopia. We enrolled patients at three sentinel hospital sites. Using RT-PCR, we screened serum samples for three arboviruses followed by serotyping and sequencing for DENV-positive samples (10.4% of samples). We detected two DENV serotypes (DENV1 and DENV3). Phylogenetic analysis identified one transmission cluster of DENV1 (genotype III major lineage A), and two clusters of DENV3 (genotype III major lineage B). The first showed close evolutionary relationship to the 2023 Italian outbreak and the second cluster to Indian isolates. Co-circulation of DENV1 and DENV3 in some regions of Ethiopia highlights the potential for severe dengue. Intensified surveillance and coordinated public health response are needed to address the threat of severe dengue outbreaks.

Molecular epidemiology of hepatitis B, hepatitis C, and HIV-1 co-infections in Ethiopia: Implications for disease burden and intervention strategies.

BACKGROUND: Hepatitis B virus (HBV) exhibits high prevalence rates within Ethiopia. The genetic diversity of HBV, marked by mixed genotype infections, may hold significant implications for the trajectory of disease and responses to treatment. Ethiopia grapples with a substantial public health challenge posed by co-infections involving HBV, hepatitis C virus (HCV), and human immunodeficiency virus 1 (HIV-1), particularly among vulnerable populations. METHODS: A comprehensive investigation into HBV, HCV, and HIV-1 co-infection was conducted. A total of 7,789 blood samples were meticulously analyzed, among which 815 exhibited HBV positivity. Among the HBV-positive samples, 630 were subjected to genotyping procedures, resulting in the identification of a prevalent trend of mixed infections characterized by HBV genotypes A/E/F (67.30%). Serological assessments were performed on 492 specimens to ascertain the presence of HCV and HIV-1 co-infections, revealing respective co-infection rates of 13.02% for HBV/HIV, 3.31% for HBV/HCV, and 2.07% for triple infection. RESULTS: The investigation revealed the intricate prevalence of co-infections in Ethiopia, notably underlining the continued transmission of viruses. The prominent occurrence of mixed HBV genotypes A/E/F suggests dynamic viral interactions and ongoing transmission pathways. These findings accentuate the necessity for targeted interventions and enhanced patient care, as co-infections carry significant clinical complexities. CONCLUSIONS: This study furnishes crucial insights into the molecular epidemiology of HBV, HCV, and HIV-1 co-infections in Ethiopia. The acquired knowledge can contribute to the advancement of strategies for clinical management and the formulation of public health interventions aimed at ameliorating the burden of viral infections within the nation.

Simplified molecular diagnosis of visceral leishmaniasis: Laboratory evaluation of miniature direct-on-blood PCR nucleic acid lateral flow immunoassay.

BACKGROUND: Diagnosis of visceral leishmaniasis (VL) in resource-limited endemic regions is currently based on serological testing with rK39 immunochromatographic tests (ICTs). However, rK39 ICT frequently has suboptimal diagnostic accuracy. Furthermore, treatment monitoring and detection of VL relapses is reliant on insensitive and highly invasive tissue aspirate microscopy. Miniature direct-on-blood PCR nucleic acid lateral flow immunoassay (mini-dbPCR-NALFIA) is an innovative and user-friendly molecular tool which does not require DNA extraction and uses a lateral flow strip for result read-out. This assay could be an interesting candidate for more reliable VL diagnosis and safer test of cure at the point of care. METHODOLOGY/PRINCIPLE FINDINGS: The performance of mini-dbPCR-NALFIA for diagnosis of VL in blood was assessed in a laboratory evaluation and compared with the accuracy of rK39 ICTs Kalazar Detect in Spain and IT LEISH in East Africa. Limit of detection of mini-dbPCR-NALFIA was 650 and 500 parasites per mL of blood for Leishmania donovani and Leishmania infantum, respectively. In 146 blood samples from VL-suspected patients from Spain, mini-dbPCR-NALFIA had a sensitivity of 95.8% and specificity 97.2%, while Kalazar Detect had a sensitivity of 71.2% and specificity of 94.5%, compared to a nested PCR reference. For a sample set from 58 VL patients, 10 malaria patients and 68 healthy controls from Ethiopia and Kenya, mini-dbPCR-NALFIA had a pooled sensitivity of 87.9% and pooled specificity of 100% using quantitative PCR as reference standard. IT LEISH sensitivity and specificity in the East African samples were 87.9% and 97.4%, respectively. CONCLUSIONS/SIGNIFICANCE: Mini-dbPCR-NALFIA is a promising tool for simplified molecular diagnosis of VL and follow-up of treated patients in blood samples. Future studies should evaluate its use in endemic, resource-limited settings, where mini-dbPCR-NALFIA may provide an accurate and versatile alternative to rK39 ICTs and aspirate microscopy.

COVID-19 vaccine reactogenicity among participants enrolled in the GENCOV study.

BACKGROUND: GENCOV is a prospective, observational cohort study of COVID-19-positive adults. Here, we characterize and compare side effects between COVID-19 vaccines and determine whether reactogenicity is exacerbated by prior SARS-CoV-2 infection. METHODS: Participants were recruited across Ontario, Canada. Participant-reported demographic and COVID-19 vaccination data were collected using a questionnaire. Multivariable logistic regression was performed to assess whether vaccine manufacturer, type, and previous SARS-CoV-2 infection are associated with reactogenicity. RESULTS: Responses were obtained from n = 554 participants. Tiredness and localized side effects were the most common reactions across vaccine doses. For most participants, side effects occurred and subsided within 1-2 days. Recipients of Moderna mRNA and AstraZeneca vector vaccines reported reactions more frequently compared to recipients of a Pfizer-BioNTech mRNA vaccine. Previous SARS-CoV-2 infection was independently associated with developing side effects. CONCLUSIONS: We provide evidence of relatively mild and short-lived reactions reported by participants who have received approved COVID-19 vaccines.

Comparison of the Diagnostic Performances of Five Different Tests in Diagnosing Visceral Leishmaniasis in an Endemic Region of Ethiopia.

The lack of accurate and feasible diagnostic tests poses a significant challenge to visceral leishmaniasis (VL) healthcare services in endemic areas. To date, various VL diagnostic tests have been or are being developed, and their diagnostic performances need to be assessed. In the present study, the diagnostic performances of rk39 RDT, the direct agglutination test (DAT), microscopy, loop-mediated isothermal amplification (LAMP), and miniature direct-on-blood polymerase chain reaction-nucleic acid lateral flow immunoassay (mini-dbPCR-NALFIA) were assessed using quantitative polymerase chain reaction (qPCR) as the reference test in an endemic region of Ethiopia. In this study, 235 suspected VL cases and 104 non-endemic healthy controls (NEHCs) were recruited. Among the suspected VL cases, 144 (61.28%) tested positive with qPCR. The sensitivities for rk39 RDT, DAT, microscopy, LAMP assay, and mini-dbPCR-NALFIA were 88.11%, 96.50%, 76.58%, 94.33%, and 95.80%, respectively. The specificities were 83.33%, 97.96%, 100%, 97.38%, and 98.92% for rk39 RDT, DAT, microscopy, LAMP assay, and mini-dbPCR-NALFIA, respectively. In conclusion, rk39 RDT and microscopy exhibited lower sensitivities, while DAT demonstrated excellent performance. LAMP and mini-dbPCR-NALFIA showed excellent performances with feasibility for implementation in remote endemic areas, although the latter requires further evaluation in such regions.

Characterizing Risk Factors for Hospitalization and Clinical Characteristics in a Cohort of COVID-19 Patients Enrolled in the GENCOV Study.

The GENCOV study aims to identify patient factors which affect COVID-19 severity and outcomes. Here, we aimed to evaluate patient characteristics, acute symptoms and their persistence, and associations with hospitalization. Participants were recruited at hospital sites across the Greater Toronto Area in Ontario, Canada. Patient-reported demographics, medical history, and COVID-19 symptoms and complications were collected through an intake survey. Regression analyses were performed to identify associations with outcomes including hospitalization and COVID-19 symptoms. In total, 966 responses were obtained from 1106 eligible participants (87% response rate) between November 2020 and May 2022. Increasing continuous age (aOR: 1.05 [95%CI: 1.01-1.08]) and BMI (aOR: 1.17 [95%CI: 1.10-1.24]), non-White/European ethnicity (aOR: 2.72 [95%CI: 1.22-6.05]), hypertension (aOR: 2.78 [95%CI: 1.22-6.34]), and infection by viral variants (aOR: 5.43 [95%CI: 1.45-20.34]) were identified as risk factors for hospitalization. Several symptoms including shortness of breath and fever were found to be more common among inpatients and tended to persist for longer durations following acute illness. Sex, age, ethnicity, BMI, vaccination status, viral strain, and underlying health conditions were associated with developing and having persistent symptoms. By improving our understanding of risk factors for severe COVID-19, our findings may guide COVID-19 patient management strategies by enabling more efficient clinical decision making.

Prognostic Value of C-Reactive Protein in SARS-CoV-2 Infection: A Simplified Biomarker of COVID-19 Severity in Northern Ethiopia.

Purpose: To evaluate the role of C-reactive protein (CRP) in predicting severe COVID-19 patients. Methods: A prospective observational cohort study was conducted from July 15 to October 28, 2020, at Kuyha COVID-19 isolation and treatment center hospital, Mekelle City, Northern Ethiopia. A total of 670 blood samples were collected serially. SARS-CoV-2 infection was confirmed by RT-PCR from nasopharyngeal swabs and CRP concentration was determined using Cobas Integra 400 Plus (Roche). Data were analyzed using STATA version 14. P-value <0.05 was considered statistically significant. Results: Overall, COVID-19 patients had significantly elevated CRP at baseline when compared to PCR-negative controls [median 11.1 (IQR: 2.0-127.8) mg/L vs 0.9 (IQR: 0.5-1.9) mg/L; p=0.0004)]. Those with severe COVID-19 clinical presentation had significantly higher median CRP levels compared to those with non-severe cases [166.1 (IQR: 48.6-332.5) mg/L vs 2.4 (IQR: 1.2-7.6) mg/L; p<0.00001)]. Moreover, COVID-19 patients exhibited higher median CRP levels at baseline [58 (IQR: 2.0-127.8) mg/L] that decreased significantly to 2.4 (IQR: 1.4-3.9) mg/L after 40 days after symptom onset (p<0.0001). Performance of CRP levels determined using ROC analysis distinguished severe from non-severe COVID-19 patients, with an AUC value of 0.83 (95% CI: 0.73-0.91; p=0.001; 77.4% sensitivity and 89.4% specificity). In multivariable analysis, CRP levels above 30 mg/L were significantly associated with an increased risk of developing severe COVID-19 for those who have higher ages and comorbidities (ARR 3.99, 95% CI: 1.35-11.82; p=0.013). Conclusion: CRP was found to be an independent determinant factor for severe COVID-19 patients. Therefore, CRP levels in COVID-19 patients in African settings may provide a simple, prompt, and inexpensive assessment of the severity status at baseline and monitoring of treatment outcomes.

HLA Variation and SARS-CoV-2 Specific Antibody Response.

Differences in SARS-CoV-2-specific immune responses have been observed between individuals following natural infection or vaccination. In addition to already known factors, such as age, sex, COVID-19 severity, comorbidity, vaccination status, hybrid immunity, and duration of infection, inter-individual variations in SARS-CoV-2 immune responses may, in part, be explained by structural differences brought about by genetic variation in the human leukocyte antigen (HLA) molecules responsible for the presentation of SARS-CoV-2 antigens to T effector cells. While dendritic cells present peptides with HLA class I molecules to CD8+ T cells to induce cytotoxic T lymphocyte responses (CTLs), they present peptides with HLA class II molecules to T follicular helper cells to induce B cell differentiation followed by memory B cell and plasma cell maturation. Plasma cells then produce SARS-CoV-2-specific antibodies. Here, we review published data linking HLA genetic variation or polymorphisms with differences in SARS-CoV-2-specific antibody responses. While there is evidence that heterogeneity in antibody response might be related to HLA variation, there are conflicting findings due in part to differences in study designs. We provide insight into why more research is needed in this area. Elucidating the genetic basis of variability in the SARS-CoV-2 immune response will help to optimize diagnostic tools and lead to the development of new vaccines and therapeutics against SARS-CoV-2 and other infectious diseases.

Case Report: Disseminated Strongloides stercoralis Presenting as an Ulcerated Gastric Mass in an HIV-1-infected Patient.

Disseminated Strongloides stercoralis is a common phenomenon among patients with immunosuppression. In this report, we present a case of disseminated Strongloides stercoralis presenting as a gastric mass in a 42-year-old male patient with a known history of HIV-1 infection and type 2 diabetes mellitus (T2DM). The patient presented with symptoms and signs suggestive of acute on chronic erosive gastritis, which included persistent vomiting. Endoscopic examination revealed a gastric mass with no signs of malignancy or dysplasia. There was noted to be chronic inflammation along with morphologic features consistent with the larvae and eggs of Strongloides nematodes in a biopsied gastric mass tissue and duodenum. The disease subsequently resulted in death despite the administration of ivermectin.

Longitudinal profile of antibody response to SARS-CoV-2 in patients with COVID-19 in a setting from Sub-Saharan Africa: A prospective longitudinal study.

BACKGROUND: Serological testing for SARS-CoV-2 plays an important role for epidemiological studies, in aiding the diagnosis of COVID-19, and assess vaccine responses. Little is known on dynamics of SARS-CoV-2 serology in African settings. Here, we aimed to characterize the longitudinal antibody response profile to SARS-CoV-2 in Ethiopia. METHODS: In this prospective study, a total of 102 PCR-confirmed COVID-19 patients were enrolled. We obtained 802 plasma samples collected serially. SARS-CoV-2 antibodies were determined using four lateral flow immune-assays (LFIAs), and an electrochemiluminescent immunoassay. We determined longitudinal antibody response to SARS-CoV-2 as well as seroconversion dynamics. RESULTS: Serological positivity rate ranged between 12%-91%, depending on timing after symptom onset. There was no difference in positivity rate between severe and non-severe COVID-19 cases. The specificity ranged between 90%-97%. Agreement between different assays ranged between 84%-92%. The estimated positive predictive value (PPV) for IgM or IgG in a scenario with seroprevalence at 5% varies from 33% to 58%. Nonetheless, when the population seroprevalence increases to 25% and 50%, there is a corresponding increases in the estimated PPVs. The estimated negative-predictive value (NPV) in a low seroprevalence scenario (5%) is high (>99%). However, the estimated NPV in a high seroprevalence scenario (50%) for IgM or IgG is reduced significantly to 80% to 85%. Overall, 28/102 (27.5%) seroconverted by one or more assays tested, within a median time of 11 (IQR: 9-15) days post symptom onset. The median seroconversion time among symptomatic cases tended to be shorter when compared to asymptomatic patients [9 (IQR: 6-11) vs. 15 (IQR: 13-21) days; p = 0.002]. Overall, seroconversion reached 100% 5.5 weeks after the onset of symptoms. Notably, of the remaining 74 COVID-19 patients included in the cohort, 64 (62.8%) were positive for antibody at the time of enrollment, and 10 (9.8%) patients failed to mount a detectable antibody response by any of the assays tested during follow-up. CONCLUSIONS: Longitudinal assessment of antibody response in African COVID-19 patients revealed heterogeneous responses. This underscores the need for a comprehensive evaluation of seroassays before implementation. Factors associated with failure to seroconvert needs further research.

Performance of rapid rk39 tests for the diagnosis of visceral leishmaniasis in Ethiopia: a systematic review and meta-analysis.

BACKGROUND: Visceral Leishmaniasis (VL) is a severely neglected disease affecting millions of people with high mortality if left untreated. In Ethiopia, the primary laboratory diagnosis of VL is by using an antigen from a 39-amino acid sequence repeat of a kinesin-related (rK39) of leishmania donovani complex (L. donovani), rapid diagnostic tests (RDT). Different rk39 RDT brands are available with very variable performance and studies from Ethiopia showed a very wide range of sensitivity and specificity. Therefore, a systematic review and meta-analysis were conducted to determine the pooled sensitivity and specificity of rk39 RDT in Ethiopia. METHOD: PUBMED, EMBASE, and other sources were searched using predefined search terms to retrieve all relevant articles from 2007 to 2020. Heterogeneity was assessed by visually inspecting summary receiver operating curves (SROC), Spearman correlation coefficient (rs), Cochran Q test statistics, inconsistency square (I2) and subgroup analysis. The presence and statistical significance of publication bias were assessed by Egger's test at p < 0.05, and all the measurements showed the presence of considerable heterogeneity. Quality assessment of diagnostic accuracy studies (QUADAS-2) checklists was used to check the qualities of the study. RESULTS: A total of 664 articles were retrieved, and of this 12 articles were included in the meta-analysis. Overall pooled sensitivity and specificity of the rk39 RDT to diagnose VL in Ethiopia were 88.0% (95% CI 86.0% to 89.0%) and 84.0% (95% CI 82.0% to 86.0%), respectively. The sensitivity and specificity of the rk39 RDT commercial test kits were DiaMed: 86.9% (95% CI 84.3% to 89.1%) and 82.2% (95% CI 79.3% to 85.0%), and InBios: 80.0% (95% CI 77.0% to 82.8%) and 97.4% (95% CI 95.0% to 98.8%), respectively. CONCLUSION: Referring to our result, rk39 RDT considered an essential rapid diagnostic test for VL diagnosis. Besides to the diagnostic accuracy, the features such as easy to perform, quick (10-20 min), cheap, equipment-free, electric and cold chain free, and result reproducibility, rk39 RDT is advisable to remains in practice as a diagnostic test at least in the remote VL endemic localities till a better test will come.

Prognostic factors and outcomes of COVID-19 cases in Ethiopia: multi-center cohort study protocol.

BACKGROUND: The coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus (SARS-CoV-2) and became pandemic after emerging in Wuhan, China, in December 2019. Several studies have been conducted to understand the key features of COVID-19 and its public health impact. However, the prognostic factors of COVID-19 are not well studied in the African setting. In this study, we aim to determine the epidemiological and clinical features of COVID-19 cases, immunological and virological courses, interaction with nutritional status, and response to treatment for COVID-19 patients in Ethiopia. METHODS: A multi-center cohort study design will be performed. Patients with confirmed COVID-19 infection admitted to selected treatment centers will be enrolled irrespective of their symptoms and followed-up for 12 months. Baseline epidemiological, clinical, laboratory and imaging data will be collected from treatment records, interviews, physical measurements, and biological samples. Follow-up data collection involves treatment and prognostic outcomes to be measured using different biomarkers and clinical parameters. Data collection will be done electronically using the Open Data Kit (ODK) software package and then exported to STATA/SPSS for analysis. Both descriptive and multivariable analyses will be performed to assess the independent determinants of the treatment outcome and prognosis to generate relevant information for informed prevention and case management. The primary outcomes of this study are death/survival and viral shedding. Secondary outcomes include epidemiological characteristics, clinical features, genetic frequency shifts (genotypic variations), and nutritional status. DISCUSSION: This is the first large prospective cohort study of patients in hospitals with COVID-19 in Ethiopia. The results will enable us to better understand the epidemiology of SARS-CoV-2 in Africa. This study will also provide useful information for effective public health measures and future pandemic preparedness and in response to outbreaks. It will also support policymakers in managing the epidemic based on scientific evidence. TRIAL REGISTRATION: The Protocol prospectively registered in ClinicalTrials.gov (NCT04584424) on 30 October, 2020.

Effect of co-infection with intestinal parasites on COVID-19 severity: A prospective observational cohort study.

Background: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection results in a spectrum of clinical presentations. Evidence from Africa indicates that significantly less COVID-19 patients suffer from serious symptoms than in the industrialized world. We and others previously postulated a partial explanation for this phenomenon, being a different, more activated immune system due to parasite infections. Here, we aimed to test this hypothesis by investigating a potential correlation of co-infection with parasites with COVID-19 severity in an endemic area in Africa. Methods: Ethiopian COVID-19 patients were enrolled and screened for intestinal parasites, between July 2020 and March 2021. The primary outcome was the proportion of patients with severe COVID-19. Ordinal logistic regression models were used to estimate the association between parasite infection, and COVID-19 severity. Models were adjusted for sex, age, residence, education level, occupation, body mass index, and comorbidities. Findings: 751 SARS-CoV-2 infected patients were enrolled, of whom 284 (37.8%) had intestinal parasitic infection. Only 27/255 (10.6%) severe COVID-19 patients were co-infected with intestinal parasites, while 257/496 (51.8%) non-severe COVID-19 patients were parasite positive (p<0.0001). Patients co-infected with parasites had lower odds of developing severe COVID-19, with an adjusted odds ratio (aOR) of 0.23 (95% CI 0.17-0.30; p<0.0001) for all parasites, aOR 0.37 ([95% CI 0.26-0.51]; p<0.0001) for protozoa, and aOR 0.26 ([95% CI 0.19-0.35]; p<0.0001) for helminths. When stratified by species, co-infection with Entamoeba spp., Hymenolepis nana, Schistosoma mansoni, and Trichuris trichiura implied lower probability of developing severe COVID-19. There were 11 deaths (1.5%), and all were among patients without parasites (p = 0.009). Interpretation: Parasite co-infection is associated with a reduced risk of severe COVID-19 in African patients. Parasite-driven immunomodulatory responses may mute hyper-inflammation associated with severe COVID-19. Funding: European and Developing Countries Clinical Trials Partnership (EDCTP) - European Union, and Joep Lange Institute (JLI), The Netherlands. Trial registration: Clinicaltrials.gov: NCT04473365.

Chronic Immune Activation and CD4+ T Cell Lymphopenia in Healthy African Individuals: Perspectives for SARS-CoV-2 Vaccine Efficacy.

Chronic immune activation has been considered as the driving force for CD4+ T cell depletion in people infected with HIV-1. Interestingly, the normal immune profile of adult HIV-negative individuals living in Africa also exhibit chronic immune activation, reminiscent of that observed in HIV-1 infected individuals. It is characterized by increased levels of soluble immune activation markers, such as the cytokines interleukin (IL)-4, IL-10, TNF-α, and cellular activation markers including HLA-DR, CD-38, CCR5, coupled with reduced naïve and increased memory cells in CD4+ and CD8+ subsets. In addition, it is accompanied by low CD4+ T cell counts when compared to Europeans. There is also evidence that mononuclear cells from African infants secrete less innate cytokines than South and North Americans and Europeans in vitro. Chronic immune activation in Africans is linked to environmental factors such as parasitic infections and could be responsible for previously observed immune hypo-responsiveness to infections and vaccines. It is unclear whether the immunogenicity and effectiveness of anti-SARS-CoV-2 vaccines will also be reduced by similar mechanisms. A review of studies investigating this phenomenon is urgently required as they should inform the design and delivery for vaccines to be used in African populations.

Utility of the Loop-Mediated Isothermal Amplification Assay for the Diagnosis of Visceral Leishmaniasis from Blood Samples in Ethiopia.

Rapid and accurate diagnosis of visceral leishmaniasis (VL) is needed to initiate prompt treatment to reduce morbidity and mortality. Here, we evaluated the performance of loop-mediated isothermal amplification (LAMP) assay for the diagnosis of VL from blood in an endemic area in Ethiopia. LAMP was positive in 117/122 confirmed VL cases and negative in 149/152 controls, resulting in a sensitivity of 95.9% (95% CI: 90.69-98.66) and a specificity of 98.0% (95% CI: 94.34-99.59), respectively. The sensitivity of the LAMP assay was 95.0% (95% CI: 88.61-98.34) in HIV-negatives and 100% (95% CI: 85.18-100.0) in HIV-positives. Compared with microscopy, LAMP detected 82/87 (94.3%, 95% CI: 87.10-98.11) of the microscopy+ cases and was negative in 11/27 (40.7%, 95% CI: 22.39-61.20) of the microscopy- cases. Compared with the rK39 serology, LAMP detected 113/120 (94.2%, 95% CI: 88.35-97.62) of the rK39+ cases and was negative in 149/154 (96.8%, 95% CI: 92.59-98.94) of the rK39- cases. However, when compared with microscopy only, rK39 detected 83/87 (95.4%, 95% CI: 88.64-98.73) of the microscopy+ cases and negative in only 12/27 (44.4%, 95% CI: 25.48-64.67) of the microscopy- cases. There was an excellent agreement between rK39 and LAMP (Kappa = 0.91, 95% CI: 0.86-0.96). Furthermore, an algorithm using rK39 followed by LAMP would yield a sensitivity of 99.2% (95%CI: 95.52-99.89) and a specificity of 98.0% (95% CI: 94.34-99.59). The findings demonstrate that LAMP assay is an accurate and rapid molecular assay for VL diagnosis, including in HIV-1 coinfected patients, in an endemic setting.

Effect of rK39 testing in guiding treatment initiation and outcome in patients with visceral leishmaniasis in Ethiopia: A prospective cohort study.

BACKGROUND: The rapid diagnostic test (RDT) rK39 is currently being used for routine diagnosis of visceral leishmaniasis (VL) in East Africa. However, continuous monitoring of the performance of the assay, in particular its impact on the clinical decision in initiating anti-leishmanial treatment and outcomes remains needed as there are concerns about the diagnostic performance of this test. METHODS: VL patients prospectively enrolled in a diagnostic trial and with rK39 RDT were included. We evaluated the effect of rK39 testing in guiding treatment initiation and outcome. On the basis of rK39 RDT test result as well as clinical case definition for VL and microscopy examination, the clinicians decide whether to initiate VL therapy or not. Poisson regression models were used to identify factors associated with a decision to initiate VL therapy. In addition, treatment outcomes of those who received VL therapy were compared to those who received non-VL treatment. RESULTS: Of 324 VL suspects enrolled, 184 (56.8%) were rK39+ and 140 (43.2%) were rK39‒. In addition, microscopy exam was done on tissue aspirates from a sub-population (140 individuals), which is 43.2% of the suspected cases, comprising of 117 (63.6%) rK39+ and only 23 (16.4%) rK39‒ cases. Of those with microscopy examination, only 87 (62.1%) were found positive. Among 184 (56.8%) patients without microscopy, 67 (36.4%) were rK39+, of whom 83 (65.9%) were positive by microscopy, 21 (16.7%) were negative by microscopy and 22 (17.5%) had no microscopy results. On the other hand, of those who did not receive VL treatment 58/189 (30.7%) were rK39+ and 131 (69.3%) were rK39‒. Of the rK39+ cases who did not receive VL therapy, only 1 (1.7%) patient was microscopy positive, 12 (20.7%) were negative and 45 (77.6%) patients had no microscopy result. Of the rK39‒ cases (n = 131) who did not receive VL treatment, 16 were microscopy negative and 115 without microscopy exams. Whereas positive rK39 result [adjusted Relative Risk (aRR) 0.69; 95% CI: 0.49-0.96, p = 0.029] and positive microscopy results (aRR 0.03; 95% CI: 0.00-0.24, p = 0.001) were independently associated with VL treatment, having confirmed diagnosis other than VL (aRR 1.64; 95% CI: 1.09-2.46, p = 0.018) was independently associated with initiation of non-VL therapy. The proportion of rK39+ patients who received non-VL treatment with no improvement outcome was significantly higher when compared to those who received VL treatment (24.1%, 95% CI: 14.62-37.16 vs. 11.9%, 95%CI: 7.26-18.93; p<0.0001). CONCLUSION: The study shows that a significant proportion of patients with rK39+ results were undertreated. Failure to do microscopy was associated with lack of improved clinical outcome. Including an additional simple point-of-care assay in the diagnostic work-up is urgently needed to correctly identify VL cases and to prevent morbidity and mortality associated with the disease.