Medical Microbiology
Permanent URI for this collection
Browse
Recent Submissions
Item Bionomics, Insecticide Susceptibility Status and Community Perception of Aedes Aegypti in Afar Region, Ethiopia(Addis Ababa Universtity, 2025) Mohammed Seid; Abebe Animut; Esayas AkliluBackground: The recent emergence and re-emergence of Aedes-transmitted diseases, such as dengue fever and chikungunya, are public health concerns in Ethiopia, including the Afar Region. Despite the frequent outbreaks of the diseases, control strategies are limited in the country. Due to the absence of licensed vaccines (except yellow fever) against Aedes-borne diseases, control of the Aedes aegypti that transmits the diseases is a priority. A successful Ae. aegypti control strategy, in turn, depends on knowledge of its behavior, insecticide susceptibility status, and vectorial role. Therefore, this research was designed to investigate the bionomics, including the spatial and temporal occurrence and larval/pupal habitat characteristics, adult resting behavior blood meal source, insecticide susceptibility, and viral infection status of Ae. aegypti. In addition, the study also assessed the community awareness about Aedes mosquitoes and associated risks to chikungunya and dengue fever. Methodology: Longitudinal study design was employed to collect adult and immature Aedes species in Awash Sebat, Awash Arba, and Werer towns of Southern Afar Regional State once per month from May 2022 to April 2023. The larvae/pupae were collected and reared to adults and identified by genus and species morphologically. The physical and chemical properties of their habitats were also characterized. Adult mosquitoes were collected using a Prokopack aspirator between 8:00 to 14:00 hrs and 15:00 to 18:00 hrs both indoors and outdoors of the house. The collected mosquitoes were sorted by sex, date of collection, collection places, and abdominal status and identified using standard morphological keys. Blood meal sources and dengue virus and chikungunya virus infection status of Ae. aegypti were determined using enzyme-linked immunosorbent assay and reverse transcriptase quantitative polymerase chain reaction, respectively. Larvae/pupae of Ae aegypti were collected and reared to test the insecticide susceptibility status of the species. Non-blood-fed, 3-5-day-old females Ae. aegypti were exposed to pyrethroid, carbamate, and organophosphate insecticide impregnated papers in tube tests following WHO standard guidelines. Knockdown rates were noted at 10-minutes interval until one hour. The mortality in Ae. aegypti was recorded 24 hrs after 60 minutes of exposure. A total of 384 respondents participated in the questionnaire survey. In addition, three focus group discussions were held to reinforce the questionnaire survey. Results: A total of 9099 Aedes larvae/pupae were collected, of which 4875 (53.6%) were from Awash Sebat, 2687 (29.5%) from Awash Arba, and 16.9% (1537) from Werer. Water holding tyres harbored the highest number of Aedes larvae/pupae followed by water storage Conclusion: Diverse types of artificial water-holding containers, including tyres, water storage drums, cement tanks, flowerpots, and discarded plastics were positive for Ae. aegypti larvae/pupae. Used tyres located at domestic sites were the most preferred Ae. aegypti larval habitats and adult resting sites in the study towns. Ae. aegypti showed resistance to bendiocarb and propoxur, suspected resistance to alpha-cypermethrin, and susceptible to deltamethrin, permethrin and pirimiphos-methyl. Health professionals, students, and merchants had more knowledge of Aedes’ larval habitats. Respondents aged 29-38 and university-educated respondents were more involved in controlling Aedes mosquito bites. Domestic water-holding containers, which serve as both immature larval habitats and adult resting sites such as used tyres and other potential larval/resting habitats management strategy is recommended in controlling Ae. aegypti abundance and hence prevention of Aedes transmitted diseases. In addition, Aedes mosquito control strategies should also be designed on the basis of the rising knowledge, risk perception, and prevention practices of the community about Aedes mosquitoes and their associated risks towards local chikungunya and dengue fever by targeting people with lower education backgrounds. Key words; Aedes aegypti, Afar Region, Attitude, Chikungunya, Dengue fever, Ethiopia, habitat characteristics, Knowledge, PracticeItem Therapeutic Effectiveness of Artemether-Lumefantrine for the Treatment of Uncomplicated Plasmodium Falciparum Malaria in Asayita Primary Hospital, Northeast Ethiopia(Addis Ababa Universtity, 2025) Alemayehu Sintayehu; Tadesse KebedeBackground: There was a global ‘missing millions’ gap between the incidence of tuberculosis (TB) cases and the notified cases. An active household contact (HHCs) investigation is one of the strategies to narrow this gap. It has the advantage of giving early diagnosis and preventive treatment to vulnerable and eligible groups. In many TB high-burden countries, only about 25% of HHCs completed TB screening, and 20–89% of eligible contacts did not adhere to TB preventive treatment. Delivering screening services near the community is important to narrow these gaps. Recognizing the transmission dynamics of Mycobacterium tuberculosis complex (MTBC) and identifying their prevalent lineages and genotypes are important in TB control and management. Objectives: This study investigated the epidemiology of active TB, TB contact screening practice, TB transmission dynamics, genetic diversity, and identified drug-resistance patterns in epidemiologically linked pulmonary TB (PTB) index cases and their HHCs in central part of Ethiopia. Methods: A cross-sectional study was carried out in 15 selected public health facilities of central part of Ethiopia from January 2022 to December 2023. The routine TB contact screening practice was assessed using one year (January, 2022 to December, 2022) data from the registration book of the study sites. Three hundred three voluntary bacteriologically confirmed PTB patients and 902 of their HHCs, without discrimination by age, were included in the study. Household contacts were screened using the World Health Organization (WHO) recommended TB symptom-based screening tool using a pre-tested questionnaire. A morning sputum sample was collected from 303 index cases and 182 HHCs having presumptive TB. Then it was examined on Xpert MTB/RIF Ultra assay, AFB microscopy (from sediment), and M. tuberculosis culture. The culture-positive 272 index cases and 13 HHC isolates were examined by phenotypic drug susceptibility test (pDST). A total of 13 index-HHC pairs and 24 randomly selected index case isolates were whole genome sequenced (WGS) using Illumina Nextseq 550. The TB-profiler and Ridom SeqSphere software were used for drug resistance and genotypic analysis. Data was entered into EPI Info version 7.0 databases and analysed using STATA V.17 software. Descriptive statistics were used to summarize the data. Chi-square test and Yates continuity correction test were used to analyse associated factors. Multivariable logistic regression analysis was done to investigate the associated vi risk factors for active TB in HHCs. Variables with a p-value A), as well as Ethionamide resistance (ethA p. Ile338Ser), which were absent from the WHO 2023 Catalogue of mutations in the Mycobacterium tuberculosis complex. Conclusion: The yield of routine TB contact investigation practice was low, and the quality of screening and adherence were suboptimal. The Home-to-home TB contact screening approach has a high active TB yield and was implementable in both rural and urban areas by mentoring and motivating the health extension workers. Scheduling convenient times and last-mile service delivery to contacts is very important to address the missed active TB cases in the community. Nearly one-third of the HHCs have discordant phenotypic drug-resistance profiles from the index patients. This study offers compelling proof that it is not advisable to treat close contacts without DST results based on the DST results of the supposed source case. Individuals residing in areas with a high burden of tuberculosis are at risk of contracting the disease from sources beyond their homes. Keywords: Active tuberculosis yield, Contact investigation, Household contacts, Index case, Transmission dynamics, Whole genome sequencingItem The Epidemiology, Oral Microbiome Signature, and Myctoxins Exposure of Esophageal Cancer Patient in Ethiopia(Addis Ababa Universtity, 2025) Girma Mulisa; Tamrat Abebe; Adane MihretBackground: There was a global ‘missing millions’ gap between the incidence of tuberculosis (TB) cases and the notified cases. An active household contact (HHCs) investigation is one of the strategies to narrow this gap. It has the advantage of giving early diagnosis and preventive treatment to vulnerable and eligible groups. In many TB high-burden countries, only about 25% of HHCs completed TB screening, and 20–89% of eligible contacts did not adhere to TB preventive treatment. Delivering screening services near the community is important to narrow these gaps. Recognizing the transmission dynamics of Mycobacterium tuberculosis complex (MTBC) and identifying their prevalent lineages and genotypes are important in TB control and management. Objectives: This study investigated the epidemiology of active TB, TB contact screening practice, TB transmission dynamics, genetic diversity, and identified drug-resistance patterns in epidemiologically linked pulmonary TB (PTB) index cases and their HHCs in central part of Ethiopia. Methods: A cross-sectional study was carried out in 15 selected public health facilities of central part of Ethiopia from January 2022 to December 2023. The routine TB contact screening practice was assessed using one year (January, 2022 to December, 2022) data from the registration book of the study sites. Three hundred three voluntary bacteriologically confirmed PTB patients and 902 of their HHCs, without discrimination by age, were included in the study. Household contacts were screened using the World Health Organization (WHO) recommended TB symptom-based screening tool using a pre-tested questionnaire. A morning sputum sample was collected from 303 index cases and 182 HHCs having presumptive TB. Then it was examined on Xpert MTB/RIF Ultra assay, AFB microscopy (from sediment), and M. tuberculosis culture. The culture-positive 272 index cases and 13 HHC isolates were examined by phenotypic drug susceptibility test (pDST). A total of 13 index-HHC pairs and 24 randomly selected index case isolates were whole genome sequenced (WGS) using Illumina Nextseq 550. The TB-profiler and Ridom SeqSphere software were used for drug resistance and genotypic analysis. Data was entered into EPI Info version 7.0 databases and analysed using STATA V.17 software. Descriptive statistics were used to summarize the data. Chi-square test and Yates continuity correction test were used to analyse associated factors. Multivariable logistic regression analysis was done to investigate the associated vi risk factors for active TB in HHCs. Variables with a p-value A), as well as Ethionamide resistance (ethA p. Ile338Ser), which were absent from the WHO 2023 Catalogue of mutations in the Mycobacterium tuberculosis complex. Conclusion: The yield of routine TB contact investigation practice was low, and the quality of screening and adherence were suboptimal. The Home-to-home TB contact screening approach has a high active TB yield and was implementable in both rural and urban areas by mentoring and motivating the health extension workers. Scheduling convenient times and last-mile service delivery to contacts is very important to address the missed active TB cases in the community. Nearly one-third of the HHCs have discordant phenotypic drug-resistance profiles from the index patients. This study offers compelling proof that it is not advisable to treat close contacts without DST results based on the DST results of the supposed source case. Individuals residing in areas with a high burden of tuberculosis are at risk of contracting the disease from sources beyond their homes. Keywords: Active tuberculosis yield, Contact investigation, Household contacts, Index case, Transmission dynamics, Whole genome sequencingItem Hepato cellular Carcinoma among Chronic Hepatitis B Patients at St. Paul's Hospital Millennium Medical College(Addis Ababa Universtity, 2025) Bethelhem Hailu; Nega BerheBackground: Liver cancer is the third leading cause of cancer death and the sixth most commonly diagnosed cancer globally. Hepatocellular carcinoma (HCC) accounts for up to 90% of liver cancer cases and chronic hepatitis B virus infection is one of the primary etiological factor. HCC presents a significant global health burden, particularly in regions with high (>8%) hepatitis B virus prevalence like Ethiopia. Objective: To estimate the incidence rate of HCC and associated factors among chronic hepatitis B patients enrolled in a longitudinal follow-up study at Saint Paul’s Hospital Millennium Medical College in Addis Ababa, Ethiopia since 2015 GC. Method: This study was nested in a prospective cohort of chronic hepatitis B patients. Data was extracted from patient records which includes demographic, clinical, radiologic and laboratory variables. The primary outcome was the development of HCC confirmed on two imaging studies. Incidence rate was calculated per 1000 person years. To identify factors associated with HCC Univariable Cox proportional hazard regression was used followed by Firth’s penalized multivariable regression, to address limited number of events and complete separation. Bootstrapping with 500 replication was then performed. Result: Out of 1291 eligible CHB patients, 23 were diagnosed with HCC during the 10 year follow up, of which those diagnosed after 12 months of follow up were considered to be incidences(n=11). The incidence rate of HCC was 1.65 per 1000 person years (95%CI: 0.81- 2.79) among all study participants and 7.07 per 1000 person years (95%CI: 3.91 – 12.76) among those with cirrhosis. Kaplan-Meier estimates showed cumulative HCC risk of 0.44% at 3 years, 0.86% at 5 years and 2.45% at 10 years. Firth’s penalized cox regression followed by bootstrap with 500 replications showed both increasing age (AHR: 1.07 per year, 95% CI: 1.03 - 1.12, P=0.000) and cirrhosis (AHR: 52.83, 95% CI: 27.25 – 102.43, P=0.000) were significantly associated with HCC. Conclusion: cirrhosis and increasing age were the strongest predictors of HCC in this 10 year cohort of CHB patients. Surveillance strategies especially targeting those with the strongest predictors are recommended to improve early detection. Key words: Chronic Hepatitis B, Hepatitis B Virus, Hepatocellular carcinomaItem The Genetic Landscape of Breast Cancer and Its Intersection With Host Immunity In Ethiopian Breast Cancer Patients(Addis Ababa Universtity, 2025) Meron Yohannes; Tamrat AbebeBackground; Breast cancer remains a leading cause of morbidity and mortality among Ethiopian women. It is a heterogeneous disease, both in its molecular characteristics and immune response. Recent evidence demonstrates the importance of incorporating immune cell composition into tumor classification, as the immune landscape plays a crucial role in determining prognosis and therapeutic response. Additionally, emerging studies suggest that tumor immune response is influenced by intratumoral microbiota, which can affect both anti- and pro-tumor immune activities. Despite these insights, there remains a significant gap in understanding the immunological landscape of Ethiopian breast cancer, its prognostic relevance, and its interaction with the mammary microbiota. Objective; This study was aimed to explore the host immune profile—both local and systemic— its role in predicting disease prognosis, and its interaction with the mammary microbiota in Ethiopian breast cancer patients. Methodology: A combination of prospective and retrospective follow-up study designs was employed to address the different study objectives. A prospective follow-up study was conducted to investigate the systemic immune repertoire of BC patients and to examine correlations between mammary microbiota and local immunity. Patient/disease characteristics at diagnosis were collected through structured, pretested questionnaires. Blood from BC patients and healthy controls were analyzed by flow cytometry to assess the phenotype and function of immune cell subpopulations. To explore microbiome-immune associations, fresh frozen tissue (FFT)/Formalin fixed paraffin embedded (FFPE) tissue was used for molecular characterization of the microbiota, by targeting 16S rRNA genes (Illumina MiSeq) and for immune gene expression analysis. Budget: The study was supported by the Susan Komen GTDR16378013 given to Martin-Luther –University Halle-Wittenberg, Medical Faculty. Keywords: Immune Phenotypes, Tumor immune microenvironment, Immune profiling, Immune escape, Mammary microbiota, Ethiopian patients, Prognostic biomarkers.Item Genetic profile, Epidemiology, and Community Knowledge of Schistosoma Haematobium in Endemic Regions of Ethiopia(Addis Ababa Universtity, 2025) Tigist Mohammed; Endakachew NibretBackground: Schistosomiasis (SCH) is a highly prevalent yet neglected tropical disease (NTD), particularly in sub-Saharan Africa (SSA), where it continues to impose a significant burden on public health and socio-economic development. Urogenital schistosomiasis (UGS), caused by S. haematobium, is a significant public health challenge in low-altitude regions of Ethiopia, including Gambella, Afar and Benishangul-Gumuz. While extensive research has focused on the epidemiology and treatment of SCH, critical gaps remain in understanding the genetic makeup of Schistosoma populations in Ethiopia and the potential hybridization events that may influence transmission dynamics and treatment outcomes. Additionally, comprehensive epidemiological data on S. haematobium infection is lacking across all age groups, particularly among pre-school age children (PSAC). This gap is compounded by limited data on community knowledge, attitudes, and practices (KAP) regarding UGS, which are essential for evaluating the effectiveness of current control measures and designing target-specific interventions. Objectives: This study aimed to investigate the genetic profile of S. haematobium and to identify hybridization events in Afar and Gambella endemic areas; assess the prevalence, intensity, and risk factors of S. haematobium infection among PSAC; evaluate community KAP in Gambella, Abobo district; and to determine the prevalence and infection intensity of S. haematobium across all age groups in the Kurmuk district of Benishangul-Gumuz. iii Methods: A community-based cross-sectional study was conducted between 2021 and 2024 to address all objectives. Multi-locus genetic analysis, targeting the mitochondrial cytochrome oxidase sub-unit I (cox1) gene and the nuclear internal transcribed spacer 2 (ITS2) regions, was performed on miracidia isolated from human urine samples using rapid diagnostic polymerase chain reaction (RD-PCR). Infection status was determined through a urine filtration microscopy (egg detection and count) and dipstick screened for hematuria. Socio-demographic data, risk factors, and outcome variables (KAP) were collected using a pretested structured questionnaire. Data were analyzed using SPSS version 23, with logistic regression and odds ratios employed to assess the strength of associations between dependent and independent variables. Results: A total of 177 miracidia were analyzed using mitochondrial cox1 and 24 miracidia were analyzed using nuclear ITS2 gene markers, revealing exclusive infection with S. haematobium and no evidence of mixed infection/hybridization with S. bovis or S. mansoni. This suggests mono-specific transmission in the Afar and Gambella regions, likely due to ecological separation, species-specific host preferences, and limited zoonotic transmission pathways. The prevalence of S. haematobium infection among PSAC in Abobo, Gambella, was 16.7%, with 20% being heavy-intensity infections. The prevalence of macro and microhematuria were 7.9% and 26.2%, respectively. The prevalence of infection was significantly higher among PSAC playing or bathing in infested water (AOR = 2.9, CI: 1.0–8.1, p=0.041) and residing in Abaro village (AOR = 4.3, CI: 1.6–11.9, p=0.005). The overall prevalence of S. haematobium infection was 34.2% in Kurmuk, Benishangul-Gumuz, with the highest prevalence among participants with Swimming habits (AOR = 2.43, CI: 1.19-4.94, p=0.023). Infection intensity varied significantly across villages, with mean egg counts (MEC) ranging from 2.39 in Ogendu (CI: 1.05-3.72) to 14.1 eggs/10 mL urine in Dulshatalo (CI: 4.98- 23.12). iv Hematuria prevalence was 39.2%, with a higher odd in Dulshatalo village (AOR = 2.64, 95% CI: 1.43-4.87, P = 0.004). Moreover, microhematuria was significantly associated with S. haematobium egg detections (P < 0.001), and a moderate agreement (Kappa = 0.57) was established with the microscope finding. The community KAP assessment in Abobo, Gambella, revealed that most participants (90.6%) had heard of UGS, and over 94% recognized at least one major symptom. However, more than half of the participants perceive (59.9%) drinking contaminated water as a primary way of transmission and understanding prevention measures (15.9% for keeping hygiene, 26.8% for refraining from using contaminated water) were limited. Over 50% did not associate urinating near water to transmission; nearly all had contact with dam/river water. Education and prior infection history were key predictors of better KAP. Conclusion: This study provided critical insights into the epidemiology and genetic profile of S. haematobium infections in endemic regions of Ethiopia, confirming the absence of mixed infection and suggesting mono-specific transmission in Afar and Gambella. The high prevalence of S. haematobium infection and heavy intensity, particularly among PSAC and older children (5–12 years), shows the public health burden of UGS in these areas. The study also identified key risk factors, such as water contact behaviors, and reveals significant gaps in community knowledge of UGS transmission and prevention despite high awareness of the disease. To address these challenges, targeted interventions are needed, including providing safe water sources, improved sanitation, and community-based health education programs to reduce risky behaviors and enhance prevention practices. Control programs should also be expanded to include PSAC, which is currently underserved but exhibits significant infection rates. Strengthening surveillance systems to monitor infection trends and genetic diversity and conducting further research to explore ecological and behavioral drivers of transmission will be v essential for designing effective, context-specific strategies to achieve sustainable SCH control in these endemic regions. Keywords: Genetic profile; Epidemiology; community based; PSAC; S.haematobium; knowledge, attitude, and practice; Afar, Gambella, Benishangul-Gumuz; EthiopiaItem Microbial Profile, Antimicrobial Resistance, and Outcome of Presumptive Meningitis Patients at the University of Gondar Comprehensive Specialized Hospital, Gondar, Northwest Ethiopia(Addis Ababa Universtity, 2025) Derso Wale; Alem AbrhaBackground: Meningitis is a life-threatening disease characterized by inflammation of the meninges. It remains a significant public health challenge, particularly in resource-limited settings where delayed presentation to healthcare facilities and comorbidities are more common. In Ethiopia, the etiology of meningitis is rarely identified, and treatment is often empirical, primarily targeting bacterial pathogens. However, this approach poses challenges due to the dynamic nature of pathogen prevalence and the increasing threat of antimicrobial resistance. Objectives: The study was conducted to assess the microbial profile, antimicrobial resistance, and outcome of presumptive meningitis patients at the University of Gondar Comprehensive Specialized Hospital, Gondar, Northwest Ethiopia, 2023. Methods: A prospective cross-sectional study was conducted from August 01 to December 31, 2023, on 195 presumptive meningitis patients at the University of Gondar Comprehensive Specialized Hospital. A cerebrospinal fluid sample was collected from each study participant and analyzed using culture and PCR techniques for the detection of bacterial and viral etiologies. The antimicrobial susceptibility of isolates was determined using the disc diffusion method. Patient outcomes were assessed using the Glasgow Outcome Scale. Descriptive statistics were employed. Bivariate and multivariable analyses were also done to identify factors associated with unfavorable patient outcomes. In all the cases, a p-value <0.05 at 95% confidence level was considered statistically significant. Results: The overall prevalence of meningitis among the 195 patients with presumptive meningitis was 114 (58.5%). Bacterial pathogens were detected in 55 (28.2%) of the study participants, which constituted 55/114 (48.2%) of the PCR-confirmed cases. E. coli was the most commonly detected pathogen 25 (21.9%), followed by H. influenzae 9 (7.9%), S. pneumoniae 6 (5.3%), N. meningitidis 4 (3.5%), S. aureus 3 (2.6%), K. pneumoniae 1 (0.9%), L. monocytogene 1 (0.9) and mixed infections involving both E. coli and K. pneumoniae 3 (2.6%). Viral pathogens were detected in 59 (30.3%) of the study participants, which constitute 59/114 (51.8%) of the PCR-identified etiologies. The most frequently detected virus was enterovirus 53 (46.5%), followed by herpes simplex virus-1 5 (4.4%), and Epstein-Barr virus 1 (0.9%). A total of 14 (7.2%) CSF samples were found positive for bacterial growth in culture. The most frequently isolated pathogens were S. pneumoniae and K. pneumoniae for 4 each, followed by E. coli and S. aureus 2 each, and H. influenzae and N. meningitidis 1 each. With regard to drug resistance, 3/4 (75%) of K. pneumoniae exhibited resistance to both trimethoprim-sulfamethoxazole and ampicillin and 2/4 (50%) to meropenem, cefotaxime, and ciprofloxacin. E. coli isolates showed 1/2 (50%) resistance to ceftazidime, tetracycline, ciprofloxacin, chloramphenicol, trimethoprim sulfamethoxazole, and ampicillin. 1/4 (25%) of S. pneumoniae isolates were resistant to erythromycin, trimethoprim-sulfamethoxazole, ciprofloxacin, and rifampin. 2/2 (100%) of the S. aureus isolates were resistant to both chloramphenicol and rifampin. Unfavorable outcomes on leaving the hospital were documented in 27/195 (13.8%) of the study participants. Glasgow Coma Scale (GCS) (AOR = 0.658, 95% CI = 0.524–0.827), the presence of comorbidity (AOR = 9.221, 95% CI = 1.580–53.815), and duration of disease onset to hospital visit (AOR = 1.604, 95% CI = 1.146–2.242) were factors independently associated with the unfavorable outcome; however, dexamethasone use does not confer benefit or harm in multivariate analysis. Conclusion: A considerably high prevalence of meningitis was detected. Viral meningitis was more prevalent than bacterial, but most presumptive meningitis patients were empirically treated for bacterial infections. High rates of antibiotic resistance among gram-negative bacterial isolates were observed. Most of the study participants experienced favorable outcomes; however, unfavorable outcomes were associated with low GCS, the presence of comorbidity, and delayed presentation to healthcare facilities. The use of dexamethasone as adjunctive treatment in presumptive meningitis patients does not confer benefit or harmItem Prevalence and Immune Correlates of Risk in Community-Acquired Meningitis: A Study at ALERT and Hawassa University Comprehensive Specialized Hospitals, Ethiopia.(Addis Ababa Universtity, 2025) Jemal Aman; Tamrat AbebeBackground: Meningitis poses a major public health challenge in sub-Saharan Africa, often triggering devastating epidemics. Although it can affect individuals of all ages, infants, children, and immunocompromised persons are at the greatest risk. In Ethiopia, meningitis continues to be a significant public health concern due to its potential for outbreaks, high mortality rates, and the considerable burden it places on the healthcare system. Objective: This study was aimed to assess the prevalence of etiologic agents causing meningitis and to evaluate immune correlates of risk among clinically suspected meningitis cases in Ethiopia. Methods: An institutional-based cross-sectional study was conducted from August 2023 to June 2024 at ALERT Comprehensive Specialized Hospital and Hawassa University Comprehensive Specialized Hospitals. A total of 201 clinically suspected meningitis cases of all age groups (except neonates) were enrolled. Cerebrospinal fluid (CSF) samples were analyzed using conventional culture, microscopy, and Conventional multiplex PCR to identify bacterial and viral pathogens. Immune correlates of risk including CD4+ T-cell counts and plasma complement levels (C5, C5a, C9, Factor D, MBL, Factor I, Properdin)- were compared among cases with bacterial meningitis, viral meningitis cases and healthy individuals. Univariate and multivariate logistic regression analyses were performed to identify significant associations. Results: The overall prevalence of bacterial meningitis detected by classical CSF culture was 5.5% (n = 11), with Klebsiella pneumoniae 3/11(1.5%) being the most frequently isolated organism. In contrast, multiplex PCR identified a substantially higher prevalence of bacterial meningitis at 36.8% (n = 74), with Escherichia coli 30/74(14.9%), Streptococcus pneumoniae 15/74(7.5%), and Neisseria meningitidis 11/74(5.5%) as the predominant pathogens. Klebsiella pneumoniae 9/74(4.5%), Streptococcus agalactiae 4/74(2%), and Listeria monocytogenes 2/74(1%) were also rarely detected. Antimicrobial resistance was alarmingly high, with 72.7% of bacterial isolates exhibiting resistance to three or more antibiotics. The overall prevalence of viral meningitis was 34.3% (n = 69), with Human Enteroviruses (28.9%) representing the most common viral etiology. Overall 65% [13/20] of patients with viral meningitis have CD4 deficiency [<200cells/mm3]. 54% [27/50] of patient with bacterial meningitis have Properdin deficiency followed by 52% [26/50] C5- Deficiency and 46% [23/50] C9- deficiency. Analysis of immune correlates revealed that deficiencies in complement components C5 (AOR = 2.50, 95% CI: 1.60–3.90, p < 0.001), C9 (AOR = 3.09, 95% CI: 1.31 7.30, p = 0.007), and Properdin (AOR = 1.80, 95% CI: 1.20–2.70, p = 0.003) were significantly associated with an increased risk of bacterial meningitis. For viral meningitis, low CD4+ T-cell levels were significantly associated with increased risk (AOR = 2.59, 95% CI: 1.60–3.90, p = 0.035). Conclusion and recommendation: In conclusion, our study confirms a significant burden of bacterial and viral meningitis in our study settings, disproportionately affecting young children. Our findings highlight specific immune correlates of risk for meningitis: deficiencies in terminal complement components (C5, C9, and Properdin) significantly increase susceptibility to bacterial meningitis, whereas low CD4+ T-cell levels are associated with a higher risk of viral meningitis. These findings highlight the urgent need for enhanced diagnostic capabilities, expanded vaccination programs, stringent antibiotic stewardship, and the implementation of immune profiling for better patient management in Ethiopia. Future research should track long-term outcomes and evaluate intervention effectiveness.Item Prevalence of Dengue and Chikungunya in Malaria-Suspected Febrile Patients: Mosquito-Borne Diseases’ Knowledge, Practices and Diagnostic Challenges in Selected Districts of Afar National Regional State, Northeast Ethiopia(Addis Ababa Universtity, 2025) Biruk Zerfu; Mengistu Legesse; Tesfu KassaDengue virus (DENV) and chikungunya virus (CHIKV) infections are an emerging burden to rising non-malarial febrile illnesses in tropical and subtropical regions, including Ethiopia. Ethiopia has experienced several significant outbreaks of arboviral diseases over the past two decades, particularly in the eastern and northeastern regions, including in the Afar National Regional State. Given the overlap in symptoms and the risk of co-infection, it is important to assess the true burden of these diseases among malaria-suspected patients during non-outbreak periods and understand community awareness and healthcare worker (HCW) challenges to improve public health intervention and strengthen healthcare response to these diseases. Hence, this study was designed to assess the prevalence of DENV and CHIKV infection in malaria-suspected febrile patients, community knowledge, attitudes, and practices (KAP) about these arboviral diseases and healthcare workers’ (HCWs) knowledge, practices and challenges in diagnosing and managing arboviral and non-malarial febrile illnesses in malaria-endemic selected districts of the Afar National Regional State, northeast Ethiopia. Methods: Between September 2022 to March 2023, a cross-sectional study was conducted among malaria-suspected febrile patients, with axillary body temperature of ≥37.5°C, attending healthcare facilities to assess the prevalence of CHIKV and DENV infection. Socio-demographic and clinical features were collected using a structured questionnaire. Five milliliters of venous blood were collected and thick and thin blood films were prepared for the diagnosis of malaria on-site. Sera were separated from remaining blood and stored appropriately. It was tested for anti-DENV and anti-CHIKV IgM and IgG antibodies using enzyme-linked immunosorbent assay (ELISA). IgM-positive sera were pooled and screened for viral RNA using in-house reverse transcription-polymerase chain reaction (RT-PCR) and a one-step multiplex RT-PCR. Those positive pools were disaggregated and retested individually to identify specific RNA-positive patient serum. To assess community KAP and HCWs’ knowledge, practices, and challenges, structured questionnaires, as well as focus group discussions (FGDs) with community members, in-depth interviews (IDIs) with frontline HCWs were conducted. Additionally, secondary data on febrile patient diagnosis and management were reviewed from public healthcare facilities to evaluate HCW diagnostic practices and trends. Quantitative data were coded and entered into EpiData 3.1, and exported and analyzed using Stata 14, with p-value < 0.05 statistically significant. Thematic analysis was applied to qualitative data. Results: A total of 411 malaria-suspected febrile patients (55.5% female, age ranged from 5-80 years, with a mean age ± SD of 27.3± 13.9) years participated in the prevalence study. Among 368 sera tested for anti-CHIKV antibodies, 176(47.8%) were IgM positive, indicating acute CHIKV infection and 23(6.3%) were IgG positive, suggesting previous exposure. Non-married patients and those having back pain were associated with positivity for acute CHIKV infection (AOR = 2.34, 95% CI: 1.14- 4.96 and AOR = 1.78, 95% CI: 1.08-2.95), respectively. Among 410 sera tested for anti-DENV IgM, 101(24.6%) were positive, indicating acute DENV infection. Of 367 sera tested for IgG, 142 (38.7%) were positive, indicating a previous exposure. Previous DENV exposure was significantly associated with being female (AOR = 2.24; 95% CI: 1.32-3.26) and employed (AOR = 1.90; 95% CI: 1.03-8.15) participants. From all 411 febrile patients, only 44 (10.7%) were positive for Plasmodium infection, mainly due to P. falciparum (84.1%). Co-infections were observed in 15.8% of patients for acute DENV-CHIKV, 4.4% for acute DENV-malaria and 4.3% for acute CHIKV-malaria among those tested for both infections. Out of 53-pooled samples positive for one or both viruses’ IgM, three were confirmed positive for DENV RNA, two of which were from DENV-CHIKV dual IgM-positive samples and the remaining one from a CHIKV IgM-positive only pooled sample by in-house RT-PCR, while two CHIKV RNA samples were identified from DENV-CHIKV dual IgM-positive pools using commercial RT-PCR methods. In the KAP study, 296 community members (mean age 34.2 ± 10.6 years; 36.8% female) and 116 FGD participants were included. While 67.3% of the community had heard of CHIK, only 44.7% recognized it as viral, 48.7% identified Aedes mosquitoes as vectors, and 16.5% knew they bite during the day. The mean KAP scores were moderate, with knowledge at 63.2%, attitudes at 60% and practices at 60%. Higher education (9-12) and student status were linked to good knowledge; age 45–59 and being single were associated with positive attitudes. Among 82 HCWs surveyed (plus 10 in IDIs), 93.9% had heard of CHIKV and DENV, with the overall mean knowledge score of 13.1 ± 3.9 (62.4%), showing a moderate knowledge level, but only 36.4% demonstrated good knowledge, while 35.1% showed moderate knowledge of the diseases. Higher knowledge was significantly associated with higher educational levels. The patient record review revealed over-diagnosis of malaria infection and low diagnosis of non-malarial febrile illness, while limited differential testing capability for arbovirus infections, inadequate training for HCWs, low community involvement and low awareness of arboviral and non-malarial mosquito-borne febrile diseases were identified as challenges. Conclusion: The seroprevalence of DENV and CHIKV infections is substantial among malaria-suspected acute febrile patients, highlighting the need for integration of arbovirus infections in regular screening, management and control of the infections in the study area. Limited awareness and diagnostic capability demand strengthening diagnostic capacity, updating fever case management and diagnosis policies, such as availing rapid test kits, serological tests (IgM tests) and affordable PCR tests for arboviral infections and scaling up HCW training and community sensitization are critical for effective arboviral and other mosquito-borne diseases control in malaria-endemic settings.Item Molecular Characterization of SARS-CoV-2 and Evaluation of Extraction-Free RT PCR Detection Methods for the Development of an Alternative and Affordable Diagnostic Tools(Addis Ababa Universtity, 2025) Getnet Hailu; Gadissa Bedada; Wondwossen AmogneBackground: SARS-CoV-2, the etiological agent of COVID-19, first emerged in Wuhan, China, and then disseminated globally at an extraordinary pace. It is classified as a single-stranded, positive-sense RNA virus within the Coronaviridae family and the Betacoronavirus genus. March 2nd, 2025, it has resulted in over 777 million confirmed cases and more than 7 million deaths worldwide. In Africa, over 12 million cases and 256,542 deaths have been reported, with Ethiopia reporting more than 500,000 cases and 7,574 deaths. The primary diagnostic method used for identifying infection is real-time reverse transcription quantitative PCR (RT-qPCR), which is recognized as the gold standard. In Ethiopia the number of SARS-CoV-2 sequences that represented the different pandemic episode and submitted to GISAID is very small compared to other African countries. In Addition, its detection was challenged by limited access to essential laboratory supplies, including RNA extraction kits, alongside the ongoing emergence of new variants. These factors highlight the continuous genomic surveillance of SARS-CoV-2 is essential for tracking of viral mutations, informs public health interventions, and aids in the development of effective diagnostics, vaccines, and therapeutic strategies; and demanding need for the development of an alternative diagnostic methods that are accurate, cost-effective, rapid, and less reliant on imported resources. Objectives: The study aimed to characterizes circulating SARS-CoV-2 variants during the fifth wave of the pandemic in Ethiopia and identify conserved gene regions through whole genome sequencing, as well as to evaluate the diagnostic performance of an extraction-free RT-qPCR method as a simplified approach for SARS-CoV-2 detection. Methods: 150 SARS-CoV-2 positive Ct-value of <30) samples were retrieved from the national reference repository of Ethiopian Public Health Institute for whole genome sequencing. These samples were primarily collected from COVID-19 suspected cases for clinical management purpose during the fifth wave of the pandemic (June to August 2022). These samples were retested, and 70 samples that maintained a Ct value ≤30 were selected for whole genome sequencing on the Illumina NextSeq 550 platform. Of the 70 sequenced samples, 63 yielded high-quality genomes that successfully passed all bioinformatics quality control assessments. These genomes were analyzed for variant classification, single-nucleotide polymorphism (SNP) distances, phylogenetic relationships, and conserved gene regions identification. The analytical workflow incorporated tools such as FastQC, Cutadapt, BWA, Samtools, Pangolin, Nextclade, SNP-dists, MAFFT, and IQ-TREE. Metadata and sequence data were analyzed using STATA v17. In addition, three hundred stored nasopharyngeal specimens (190 SARS-CoV-2 positive and 110 negative) were randomly selected from the national repository. The samples were processed using the standard RNA extraction kits (used as the gold standard) and an extraction-free method that involved dilution and heating (EFDH). In the EFDH protocol, samples were diluted at a 1:2 ratio with RNase-free water and subjected to heating at 72°C for 15 minutes. Then, after master mixing, RT-qPCR was performed using DAAN kits targeting the N and ORF1ab genes by the ABI 7500 platform. The results were stratified by viral load: Ct <20, 20–35, and >35 and diagnostic performance was analyzed by using R-Studio and STATA v17.Simultaneously, 150 SARS-CoV-2 positive samples (Ct ≤30) collected during the fifth wave of the pandemic in Ethiopia for genomic analysis. Results: Among the 70 samples sequenced, 63 met the quality control criteria, exhibiting a genome completeness of 99.8%. The Omicron variant was the predominant strain, accounting for 97% of the samples, while the Delta variant was present in 3% of the samples. Phylogenetic analysis revealed five principal clades. On average, 69 single-nucleotide variations (SNVs) were identified per genome. Mutations were detected within the S, M, and N genes, as well as in nonstructural proteins, with notable deletions observed in the S gene. Additional mutations were identified in PLpro and 3CLpro, and the transition/transversion (Ti/Tv) ratio was calculated to be 1.97. The sequences from Ethiopian samples exhibited low divergence (9.78×10⁴) in comparison to global sequences (1.54×10³). Two conserved regions within the nucleocapsid (N) gene were identified, encompassing amino acids 45–180 and 247–364. Three linear B-cell epitopes, residues 56–57, 160–180, and 255–275, were located within these regions. These epitopes displayed strong antigenic properties, low variability, and surface accessibility, indicating their potential as targets for future diagnostic applications.The EFDH method demonstrated a sensitivity of 86% and a specificity of 100% when compared to the standard RT-qPCR assay. It accurately identified 163 out of 190 positive samples and all 110 negative samples. The average Ct value recorded for EFDH was 27.7, in contrast to 24.1 for the standard method. A strong correlation between the two methodologies was observed (R² = 0.96, p = 0.001). EFDH exhibited 100% sensitivity for samples with Ct values less than 20 and 98% sensitivity for those with Ct values ranging from 20 to 35. However, sensitivity significantly decreased for samples with Ct values greater than 35. The overall diagnostic accuracy of the EFDH method was determined to be 91%, with a kappa value of 0.82 (p < 0.001), indicating a strong agreement with the standard method for samples with high viral loads. Conclusions: The genomic analyses revealed that the Omicron variant was the predominant strain during Ethiopia's fifth wave, suggesting ongoing community transmission and the imperative for sustained surveillance efforts. The nucleocapsid protein was validated as a stable and immunogenic diagnostic target, with identified epitope candidates facilitating future diagnostic advancements. The integration of genomic and structural data may enhance the identification of reliable diagnostic markers in the context of viral evolution. The EFDH method exhibited high sensitivity and specificity for the detection of SARS-CoV-2 in samples with elevated viral loads, demonstrating a strong correlation with the established RT-qPCR method. Nevertheless, its diminished sensitivity in cases of low viral load underscores the necessity for confirmatory testing. EFDH presents an alternative and cost-effective that suitable for resource-limited countries.Item Study on the Bionomics and Pathogen Infection Status of Aedes Mosquitoes in Metema District, Northwest Ethiopia(Addis Ababa University, 2025-10-10) Wondmeneh Jemberie; Abebe Animut; Sisay DugassaBackground: Female Aedes Species Transmit Viral Diseases Such as Yellow Fever, Dengue Fever, Chikungunya and Zika in Humans. Dengue Fever and Chikungunya Have Been Reported in Metema District, Northwest, Ethiopia However, Entomological Evidence Scares to Guide Vectors Control Strategies. Therefore, This Study Examined the Physicochemical Characteristics of Larval Breeding Habitats, Biting Time, Resting Habitats, And Wolbachia and Viral Infectious State of Aedes Mosquitoes. Methodology: Entomological Studies Were Conducted in Metema-Yohannes, Kokit and Gend-Wuha Towns, Metema Distinct, Northwestern Ethiopia from January 2022-December, 2023. A Longitudinal Entomological Study Was Carried Out on the Physico-Chemical Characteristics of Larvae/Pupae Habitats, Adult Biting Hours, Resting Habit and Cross Section Study Was Carried Out Viral and Wolbachia Strains Infection Status of Aedes Aegypti. Physical Characteristics of Larvae/Pupae Habitat Were Observed in Situ and Chemical Composition of Habitat’s Water Sample Was Analyzed Under Laboratory Condition. Aedes Mosquitoes Were Identified to Species Under Microscope Using a Taxonomic Key and Confirmed Using Molecular Technique Based on Mitochondrial Cytochrome Oxidase Subunit -1 (COI-1) And Internal Transcribed Spacer -1 & 2 (ITS-1 & 2) Genes. Biting Hour of Adult Ae. Aegypti And Ae. Vittatus Was Deterred By CDC-LT Collection from Indoor and Outdoor Each Hour For 24hrs And Their Host Seeking Behavior Was Determined by Collecting Them from Human (Indoor & Outdoor), Cattle, Goat, Sheep and Donkey Shelters Overnight. Adult Aedes Species Were Collected from Their Resting Places by Prokopenko Aspirators, Pyrethrum Spray Catches and Mouth Aspirator. Dengue Virus, Chikungunya Viruse and Zika Viruses Were Detected by SYBR Green RT-PCR And Also Wmel Strain Wolbachia Was Detected by Conventional RT-PCR. Statistical Method Used to Data Analysis Was Considered Significant When to P < 0.05. Results: Aedes Larvae/Pupae Were Abundant in Metema-Yohannes, Kokit and Genda-Wuha Towns During Wet Seasons. Breteau, House, And Container Indices Exceeded the World Health Organization (WHO) Risk Levels for Arboviral Diseases. Aedes Larvae/Pupae Density Increased in Habitat with High Water Temperature, Total Hardness, Sulphate, And with the Decrease Distance Between Households and Habitat. Adult Ae. Aegypti Larvae/Pupae Were the Most Abundant (56.77%; 5106/8993) Followed by Aedes Vittatus (37.25%; 3350/8993), Aedes Communis (2.39%; 215/8993), Aedes Opok (0.66%; 60/8993) And Aedes Albopictus (0.26; 24/8993). The Mean Ranks of Ae. Aegypti Emerged from the Larvae/Pupae of Artificial, Algae Positive and Tadpole Free Habitats Were Analyzed by Mann-Whitney U Test. Results Were Considered Significant When P < 0.05 Higher Than the Corresponding Results from the Natural, Algae Free, And Tadpole Infested, respectively. Adults Of Ae. Vittatus Followed a Similar Trend. The Mean Number of Ae. Aegypti Emerged from Larvae/Pupae of Habitats Closer to Human Habitations, Not Exposed to Sunlight, Free of Emergent Vegetation and Tyre Substrates Were Analyzed by Kruskal Wallis H Test. Results Were Considered Significantly (P<0.001) Higher Than from Habitats with No These Attributes. Aedes Aegypti And Ae. Vittatus Exhibited Two Peak Biting Rates Were Observed Indoor and Outdoor at Sunrise and Sunset. Both Ae. Aegypti And Ae. Vittatus Was Abundantly Collected from Nearby Human Sleeping Arrangements Than from the Animal Shelters Like Cattle, Sheep, Goats, And Donkeys. The Highest Proportions of Ae. Aegypti (91.21%) And Ae. Vittatus (89.87%) Was Unfed. A Total Of 4,054 Aedes Mosquitoes Were Collected That Belonged to Ae. Aegypti (3026; 58.49%), Ae. Vittatus (N = 723; 29.08%), Ae. Communis (N= 186; 7.24%), Ae Albopictus (N = 77; 3.30%), And Ae. Opok (N = 42; 1.88%). A Greater Number of Aedes Species Were Collected During the Wet Season (June to September) Than During the Dry (October to May), 2022-2023. Ae. Aegypti And Ae. Vittatus Occurred in Metema-Yohannes, Kokit, And Genda-Wuha, Aedes Communis in Genda-Wuha and Kokit, But Ae. Albopictus And Ae. Opok Only in Metema-Yohannes. Large Numbers of Ae. Aegypti, Ae. Vittatus And Ae. Communis Was Collected Resting in Cattle Shade, While Ae. Albopictus Within Larvae Hatching Containers but Ae. Opok Was Collected from Inside Bridge. The Wmel Strain of Wolbachia Was Detected in Both the First and Second Generations of Aedes Aegypti Under Laboratory Conditions, Indicating Successful Transovarian Transmission. Additionally, All Ae. Aegypti Pools Were Tested Negative For DENV, CHIKV And ZIKV. Conclusion: The Predominant Aedes Species Identified in the Study Area Was Ae. Aegypti, Followed by Ae. Vittatus. Other Aedes Species Was Observed in the Towns Included Ae. Communis, Ae. Albopictus, And Ae. Opok. Larvae And Pupae of Ae. Aegypti, Ae. Vittatus, And Ae. Albopictus Were Primarily Found in Discarded Tyres, While Ae. Communis Was Associated with Metal Containers, And Ae. Opok With Tree Holes. Peak Biting Activity for Ae. Aegypti And Ae. Vittatus Occurred During the Morning and Early Night Hours. The Resting Habitats of Aedes Species Varied with Seasonal Changes. The Pathogenic Infection Status of Ae. Aegypti May Be Influenced by Natural Suppressive Agents. Findings From This Study Can Serve as A Baseline for Implementing Targeted Aedes Mosquitoes Control Programs, Particularly in Managing Larval and Pupal Breeding Containers Within the Study Towns. Additionally, Control Efforts Should Include Community Engagement on the Elimination of Common Larval and Pupal Breeding Habitats.Item Molecular Epidemiology and Genotypic Characterization of Rotavirus Among Children Under Five Years Age with Acute Diarrheal Illness in Addis Ababa, Ethiopia(Addis Ababa University, 2025-08-04) Gemechis Dejene; Daniel Asrat; Amha MekashaAbstract Introduction: In Children Under Five Years, Diarrhea Is the Second Most Common Cause of Mortality Worldwide. Various Enteric Pathogens Are Responsible for Diarrhea, Including Viruses, Bacteria, And Parasites. Among The Viral Etiologies, Rotavirus Is the Leading Cause of Diarrheal Illness in Children. The Rotavirus-Associated Diarrheal Illness Is Highest in Sub Saharan Africa, South Asia, And Southeast Asian Countries. Rotavirus A Is Categorized in to Several G And P Genotypes Based on Vp7 and Vp4, respectively. The Most Common Genotypes Associated with Human Infection Globally Are G1p [8], G2p [4], G3p [8], G4p [8], G9p [8] And G12p [8]. Objectives: To Determine Molecular Epidemiology and Identify Circulating Genotypes of Rotavirus Among Children Under Five Years with Acute Diarrheal Illness in Selected Health Facilities in Addis Ababa, Ethiopia. Methods: A Cross-Sectional Study Design Was Conducted in Selected Health Facilities in Addis Ababa, Ethiopia, From September 2023 to January 2025. A Total Of 200 Children with Acute Diarrheal Illness Were Recruited Using a Convenience Sampling Technique. After Obtaining Consent, Caregivers Were Interviewed with a Structured, Pre-Validated Questionnaire for Sociodemographic Status, and 5 Ml of Stool Specimens Were Collected from the Study Participants. The Stools Were Tested for Rotavirus Using the Taqman Array Card -Real Time Polymerase Chain Reaction, Including Genotype Identification. Data Was Entered into Epidata Version 4.7 And Analyzed Using Spss Version 25 Software. The Results Were Presented Using Tables and Interpreted Using Chi-Square and Odds Ratio. Results: Of The 200 Stool Specimens Collected from the Children, Rotavirus Was Detected In 45 (22.5%) Samples. Thirteen G-P Genotype Combinations Have Been Identified, G1p [4], G2p [4], G3p [8], G8p [4], G9[P4], G9p [8], G2p [6], G12p [8], G10p [8], G4p [8], G1p [8], G8p [8], And G8p [6] Are the Circulating Genotypes Among the Population. Out Of These, Three New Genotypes, G8p [4], G10p [8], And G4p [8] Were Detected for The First Time in Ethiopia. Factors Associated with Rotavirus Diarrhea Were Analyzed. Generally, Only Two Factors (Vomiting and Vaccination Status) Were Statistically Significantly Associated with Rotavirus Positivity; While Other Child Related Associated Factors Were Not Significantly Correlated with Rotavirus Infection. Conclusion: This Study Provides an Important Insight into the Updated Prevalence, Circulating Genotypes and the Emergence of New Genotypes of Rotavirus Among the Population. These Findings Underline That Despite the Presence of the Rotarix Vaccine in the Country, Rotavirus Remains a Significant Etiologic Agent for Acute Gastroenteritis. Therefore, Continuous Rotavirus Surveillance and Implementation of Prevention and Control Strategies Are Crucial to Reduce the Burden of Rotavirus-Related Diarrheal Illness.Item Genotype Distribution of Human Papillomaviruses & HPV E6/E7 RNA Test for the Detection of High-Grade Cervical Intraepithelial Neoplasia (CIN2+) Among Gynecology Complaints in Northwest Ethiopia(Addis Ababa University, 2023-07) Derbie Awoke; Abebe Tamrat ; Woldeamanuel YimtubezinashBackground: In developing nations, cervical cancer (CC) is the main cause of cancer-related fatalities in women due to the absence of well-established vaccination and screening programs. Exploring the best triage test for women with cervical abnormalities is a timely area of research to advance cervical screening and management. Further, the distinct proportional impact of each HR-HPV on the distribution of cervical lesions varies geographically. There is a shortage of data regarding the clinical value of high-risk human papillomaviruses (HR-HPV) E6/E7 mRNA test and their molecular epidemiology in cervical samples from Ethiopia, particularly in the current study area. Moreover, despite the fact that HR-HPV infection is an essential biological cause of CC, other socio-demographic factors are not well studied in the nation. Therefore, this study aimed to fill these data gaps. Objectives: The aim of the study was to determine the HPV genotype involved in cervical lesions, to evaluate the clinical use of HR-HPV E6/E7 mRNA for the early detection of CIN2+, and to explore factors associated with it among gynecology complaints in northwest Ethiopia. Methods: Between March 2019 and October 2021, a cross-sectional study was carried out at Felege Hiwot Compressive Specialized Hospital (FHCSH). Among women who visited the hospital for gynecological examination, those who were eligible for visual inspection (VIA) based screening were included. Cervical punch samples were obtained by a gynecologist for histological analysis. Cervical swabs collected and analyzed for HR-HPV DNA and HPV E6/E7 mRNA using the Abbott Alinity m system and real-time PCR, respectively at the Institute of Virology, Leipzig University, Germany. Demographic and gynecologic-related history were collected using a structured questionnaire. The distribution and frequency of HR HPVs described using descriptive statistics. Histology was used as the reference test to determine how well the E6/E7 mRNA detected CIN2+. Results: Of the 355 study participants (aged 30 to 80 years), more than half, 211 (59.4%), were unaware of CC, and their previous cervical screening practice was approximately 25%. Cervical biopsies from 41.8% (140/335; 95% CI: 36.6-47.1%) participants were diagnosed as cancer. The proportion of HR-HPV was 53%(188/355; 95%CI: 47.8-58.1%), with 13 different genotypes identified. HPV16 was predominant at 50.4% (95%CI: 29.4-39.2%), followed by HPV31 (9.7%), HPV33 (8.5%), HPV39 and HPV68 (5.8% each), and HPV18 at 4.7%. The iii E6/E7 mRNA test was positive in 35.8% (127/355; 95%CI: 30.0-40.9) of cases for HPV16, 16 & 45. The proportion of positive HPV DNA test results for these three HR-HPVS was 42% (149/355). The total agreement of DNA and mRNA tests in the detection of these HPVs was at 92.7% (95%CI: 89.5-94.9) with a kappa value of 0.821. HPV16, at 108 (85%), was the most common genotype expressing E6/E7 mRNA. The mRNA assay had sensitivity, specificity, positive and negative predictive values (PPV and NPV) of 65.2% (95%CI: 57.5-72.2%), 90% (95%CI: 84.6-93.4%), 85.8% (95%CI: 78.5-91.0%), & 73.6% (95%CI: 67.2-79.1%), respectively for detecting histologically confirmed CIN2+. Specifically, the sensitivity and specificity of this assay in the detection of CIN2+ were 92.7% & 47%, respectively among HPV16, 18, & 45 DNA-positive cases. Likewise, the analytical sensitivity and specificity of the HPV-DNA test were 84.8% & 74.1%, respectively. CC increased steadily with participant age, with women older than 50 years about four times more likely to develop CIN2+ (AOR: 3.68 95%CI: 1.75-7.72, p < 0.001). Similarly, no cervical screening in the past five years (AOR: 2.04; 95%CI: 1.04-4004; p = 0.038), infection with HR-HPVs (AOR: 5.28; 95%CI: 2.66-10.47; p < 0.001) and tested positive for E6/E7 mRNA (AOR: 5.78; 95%CI: 2.73-12.24, p < 0.001) were statistically associated with CIN2+. Conclusions: CC is still a significant issue for women's health in northwest Ethiopia that requires evidence-based interventions. The E6/E7 mRNA test and the HPV DNA test demonstrated good agreement and showed better diagnostic relevance in detecting CIN2+. Therefore, the test can be considered for colposcopy and biopsy triage. In particular, the mRNA test may be regarded as a potential triage for women who are HPV-positive , mainly in regions with a shortage of pathologists and colposcopy facilities. Vaccination and future HPV-based screening methods in Ethiopia should consider the important HR-HPV genotypes identified in such studies. To better assess the HPVs circulating in northwestern Ethiopia, community-based surveys should be conducted. Likewise, to optimize the E6/E7 mRNA analytical sensitivity and specificity , large-scale studies targeting major HR-HPVs should be considered. Finally, in accordance with the WHO recommendation women who are eligible for cervical screening need to be screened with a high-precision test, including HPV-based tests. Keywords: Cervical cancer, CIN2+, HPV E6/E7 mRNA, HR-HPV DNA, northwest EthiopiaItem Burden of Methicillin Resistant Staphylococcus Aureus (MRSA) and Associated Factors at TASH: Evidence from Colonization of Health Care Workers, Administrative Staff, Patients and Selected Inanimate Objects.(Addis Ababa University, 2023-12) Tullu Kassu ; Woldeamanue Yimtubeznashl; Enquselassie Fikre; Price James; Davey GailBackground: Methicillin-resistant Staphylococcus aureus (MRSA) are multidrug-resistant bacteria that are carried harmlessly by humans and also cause severe life-threatening infections in community and healthcare settings worldwide. MRSA colonization of hospitalized patients and healthcare workers (HCWs) is a risk factor for transmission of infection. Furthermore, inanimate objects, such as Mobile phones, hospital gowns, and other fomites can serve as a reservoir for MRSA. Good knowledge, attitudes, and practices (KAP) of HCWs towards MRSA sources, colonization, and transmission is a key strategy for the control of MRSA. In Ethiopia, MRSA is a public health concern and observed rates of MRSA infections are reported from different parts of the country but molecular based data are very scarce or absent. Objectives: This study aimed to determine the burden and associated factors for MRSA colonization among HCWs, administrative staff, patients and selected inanimate objects at Tikur Anbessa Specialized Hospital (TASH), Addis Ababa, Ethiopia. Methods: A prospective and retrospective cross-sectional study was conducted between June 2018 to September, 2019 that included 588 HCWs and 468 administrative staff. Nasal swabs and swabs from their mobile phones were collected. Single swab from gown of each HCW was sampled. Samples from inanimate objects from the hospital were also analysed for MRSA. Perceived knowledge, attitudes, and practice (KAP) of HCWs were assessed using a pretested structured questionnaire. In addition, 170 S. aureus stored isolates were included mainly for molecular testing. All swabs samples were cultured on Mannitol salt agar and / or blood agar for growth and identification of S. aureus and MRSA were made by standard biochemical tests and cefoxitin disc methods respectively. Antimicrobial testing was performed according to CLSI breakpoints and isolates were further tested for MecA , PVL detection, Spa and MLST typing and WGS. Data were analysed using SPSS version 20.0 and statistical testing (Chi-square or Fisher’s exact test for categorical variables) was used to assess the difference between MRSA colonization rate among different groups, socio-demographic factors, and KAP level. P-value less than 0.05 was taken Statistically significant Results: A total of 588 HCWs were included in this study and 58.4 % of them were female. The mean age and standard deviation of HCWs were 29.13 ± 6.66 years, The majority of HCWs were nurses by profession accounting for 49.1 % (289/588) followed by Medical doctors (28.4 % ) [167/588]. Among 468 administrative staff , 64.1 % of them were females. The rate of S.aureus isolates were 16.32 % (96/588) and 8.97 % (42/468) for nasal swabs and 6.46 % (38/588) and 3,85 % (18/468) for mobile phone contamination for HCWs and administrative staff respectively . For HCWs group the rate of S. aureus isolation from gowns was 7.48 % (44/588). Overall, 31 isolates (13.02 %) were sensitive to all antibiotics tested. All isolates were sensitive to rifampicin. About 10 % of isolates including all MRSA isolates were vancomycin sensitive using Van A and Van B genes detection methods. Furthermore ,237 isolates were resistant to at least for one drug. Overall there was a significant difference between drug resistance among MRSA and MSSA isolates (P-value < 0.05). In this study, 2.72 % (16/588) and 1.3 % (6/468) mobile phones of HCWs and administrative staff were contaminated by MRSA respectively which are more frequent for nurses and doctors. Importantly ,2.89 % (17/588) of HCWs gowns were contaminated with MRSA and slightly higher among female HCWs (p-value >0.05). The proportion of MRSA from stored isolate of mother-child pair and diabetic patients was 0.24 % (2/846). While 21.73 % (20/92) stored isolates from clinical samples were MRSA. Overall 53 % (178/336) of S.aureus isolates obtained from nasal, clinical, mobile, and HCW’s gowns were PVL positive. A significant difference was also seen between pvl status and source of S.aureus isolates Key Words: S.aureus , MRSA, TASH, MLST, WGS, Spa typing, HCWsItem The Comparison of Xpert® Xpress with Cobas® SARS-CoV-2 Assay on Individual and Pooled Nasophary Neal Swabs and Whole Genome Sequencing of SARS-CoV-2 Samples Collected in Selected Hospitals of Addis Ababa, Ethiopia.(Addis Ababa University, 2023-07) Zerihun Betselot; Abegaz Woldaregay ; Gebre Selassie; Kebede Habtamu ; Alemu AyinalemBackground: Xpert® Xpress and Cobas® are assays offering rapid test for the detection of SARS-CoV-2. Pooled testing allows for continued testing even when supplies are relatively scarce. Whole genome sequencing of SARS-CoV-2 refers to the process of determining the complete genetic sequence of the virus, providing a detailed and comprehensive analysis of its entire genome. The agreement between assays for SARS-CoV-2 detection from individual and pooled samples is uncertain in Ethiopia. Moreover, there is limited availability of whole genome sequencing data for SARS-CoV-2, leading to insufficient information regarding the circulating variants, lineages, and mutations. Objective: The study aimed to compare the performance of Xpert® Xpress with Cobas® SARS CoV-2 assays for SARS-CoV-2 detection from individual and pooled nasopharyngeal swabs and whole genome sequencing of SARS-CoV-2 to identify lineages, sub-lineages and mutations for nucleocapsid and envelop gene, in Ethiopia. Methods: A cross-sectional study design was utilized on individuals suspected of having COVID-19 in seven selected hospitals in Addis Ababa, Ethiopia, between June 25, and July 20, 2022. The samples were tested for SARS-CoV-2 using the GeneXpert® and Cobas 8800®. A total of fifty samples that tested positive with the Xpert® were chosen and divided into four groups according to their CT-value and then pooled with negative samples and subjected to testing using both assays. Positive samples with a CT-value of ≤30 were selected for sequencing. The CovidSeq Assay was used to prepare sequencing libraries, which were then sequenced on the Illumina Miseq. Data were entered into Epi data version 3.1 and exported to SPSS version 28 for statistical analysis. Analysis was performed by using descriptive statistics, pearson correlation, and cohen’s kappa. Nextflow work flow manager and different bioinformatics tools were used for the analysis of variants, lineages, and mutations. Results: In this study, a total of 440 nasopharyngeal swabs were collected, and the overall rate of positivity for SARS-CoV-2 was found to be 100 (22.73%). The overall agreement between the assays from individual samples was 91.73(95% CI, 89.99% to 92.66 %, κ=0.715) indicating a substantial level of agreement. The Xpert® assay showed a high positive percent agreement (98.25%) and negative percent agreement (90.71%). The overall agreement between the assays from pooled samples was 89%, (95% CI, 86.31% to 91.52% κ=0.552). All 16 sequenced samples were identified as the Omicron variant, with ten samples assigned to clade 22A. The consensus XII sequence length ranged from 29,578 to 29,903, with a GC% ranging from 33.1% to 39.0%. The average length of amino acids in the sequences was 9519. The pairwise identity between the samples and the reference genome ranged from 98.6% to 99.9%. Several amino acid changes were observed in the sequenced samples, including P13L, P151S, N E31del, N R32del, N S33del, N P151S, N R203K, N G204R, and N S413R on the N genes. Likewise, there was one common mutation at position 9 in the E gene, E31del. Conclusion: The Xpert® performed better than the Cobas® assay in detecting positive results due to its ability to detect low viral load cases, utilizing the more sensitive nucleocapsid gene instead of the ORF1a gene used in the Cobas® assay. Confirming presumptive positive results by different methods and using multiple molecular assays targeting different genes is important to ensure high sensitivity in detecting RNA. Pooling up to six samples was possible regardless of viral load and pool size for Xpert®. But viral load effect was significant in all pool size of Cobas® assay. The agreement on individual samples indicated that both assays can be used interchangeably for diagnostic purposes. Furthermore, there were variations in amino acid length, the presence of both unique and existing mutations, and variations in mutation frequencies among the analyzed sequences. Key words: Agreement, Cobas® SARS-CoV-2 assay, COVID-19, Xpert® Xpress SARS-CoV-2 assay, Pooling, SARS-COV-2, WGSItem Human Papillomavirus Genotype Distribution, Persistence, Clearance, and Characterizing Cervicovaginal Microbiota: A Population- Based Follow up Study(Addis Ababa University, 2023-04) Endallew Brhanu ; AbebeTamrat ; Kaufmann Andreas ; Kantelhardt Eva ; Mihret AdaneBackground: Human papillomaviruses (HPVs) are a group of small, non-enveloped, naked icosahedral (55nm) viruses that can cause cervical cancer (CC) and other cancers. Cervical cancer is by far the most common HPV-related disease, and it is the second leading cause of morbidity and mortality from all cancers in Ethiopian women. Persistent infection with hr-HPVs and progression to precancerous lesions are the most important steps in the carcinogenesis process. However, most infections are transient and rarely persist implying development of CC is a multifactorial and step by step process and may require other co-factors like cervicovaginal microbiome within the local microenvironment for its development. Objective: To determine the prevalence of HPV infection, genotype distribution, the persistence and clearance rates within two years and compare the performance of different HPV tests. Furthermore, it aimed to characterize the cervicovaginal microbiota in women with premalignant dysplasia or invasive cervical cancer compared with that of healthy women. Methods: The study was conducted in two cohorts; a population-based cohort from rural women in Butajira, south-central Ethiopia and women attending gynaecological clinic at Tikur Anbessa Specialized Hospital from October 2017 to February 2020. From Butajira, a total of 893 samples were tested at baseline. A self-sampling brush (Evalyn Brush®, RoversOss The Netherlands) was used for cervical specimen collection and HPV testing was performed using multiplexed genotyping (MPG) by BSGP5+/6+ PCR with Luminex read out. Follow-up testing was done at 6 and 24 months for baseline hr-HPV positive women. Moreover, three HPV DNA testing assays (MPG-Luminex Assay, Anyplex II HPV HR Detection, and EUROArray HPV) were compared and the analytical sensitivity and specificity of the assays in detecting hr-HPV infections was computed. At Tikur Anbessa Specialized Hospital, cervicovaginal microbiota of 120 women was characterised using the 16S rRNA cervical microbiome sequencing. Shannon and Simpson diversity indexes were used to evaluate alpha diversity. Beta diversity was examined using principal coordinate analysis (PCoA) of unweighted Unifrac distances. Results: At baseline screening, the population-based HPV positivity rate was 23.2% (95% CI: 23.54‐22.86%), of these 20.5% (95% CI=20.79‐20.21), and 10.3% (95% CI=10.52‐10.08) women were hr‐ and lr‐ HPV positives, respectively. Age‐specific hr‐HPV infection peaked in the agegroup 30‐34 years old (58.6%) and decreased in 35‐39, 40‐44, and 45‐49 years to 20.4%, 4.5% and 3.8% respectively. The top five prevalent hr‐HPV genotypes were HPV16 (57.1%), 35 (20.3%), 52 (15.8%), 31 (14.1%), and 45 (9.6%) in the Butajira district. hr-HPV infection clearance was observed in 70 women (73.7%) within 6 months and among 77 women (84.6%) within 2 years. In the control women (negatives at baseline), the hr-HPV incidence was 4.1%.Item Molecular Detection of Virulence and Antimicrobial Resistance Genes of Streptococcus Pyogenes Isolated From the Oropharynx of Febrile Patients in Adama and Wonji, Ethiopia.(Addis Ababa University, 2023-08) Lemi Haile ; Negash Abel ; Mihret Adane; Adane KelemeworkBackground: Streptococcus pyogenes also called Group A Streptococcus (GAS) is a significant human pathogen that causes several diseases including pharyngitis, skin infections, rheumatic fever and rheumatic heart disease. GAS produces a variety of virulence factors such as toxins/superantigens, proteases, DNases. Detection of virulence genes can be used to determine the pathogenic potential of a strain. Although GAS is generally thought to be susceptible to β-lactams, an increasing trend of treatment failures and resistance to macrolides and tetracyclines have been observed. There is a paucity of data on the molecular epidemiology of virulence factors and antibiotic-resistance of S. pyogenes in Ethiopia. Objective: To assess virulence factors, antimicrobial susceptibility profile and drug resistance genes of oropharyngeal S. pyogenes isolated from febrile patients. Method: A cross-sectional study was conducted on 201 S. pyogenes isolated from the oropharynx of febrile patients in Adama and Wonji, Oromiya, Ethiopia. Standard microbiological procedures were used for bacterial growth and identification. Bacterial DNA was extracted using an alkaline lysis buffer. Multiplex PCR was conducted to identify virulence factors for the production of superantigens (SpeA, SpeC, SpeG, SpeH, SpeI, SpeJ, SpeK, SpeL, SpeM, SmeZ and ssa); proteases (SpeB, spyCEP, scpA, Mac and sic); and DNases (sdaB, sdaD, sdc and spd3), erythromycin resistance genes [mef(A), erm(B) and erm(A)-TR] and tetracycline resistance genes [tet(M), tet(O), tet(K)and tet(L)]. SPSS version 27 was used for statistical analysis. Majority of the results were summarized by frequency and percentage distributions. Result: This study was performed on 201 S. pyogenes isolates. We investigated the presence of 20 different GAS virulence factors and identified 18 of them with variable proportions. Relatively higher frequencies of Mac (24.9%), SmeZ (16.9%), scpA (15.4%), sic (12.9%), SpeK (12.4%) and SpeI (10.0%) genes were identified. Our antibiotic susceptibility testing showed that 10% of the GAS isolates were non susceptible to penicillin. High resistance rate to tetracycline (54.2%), quinupristin-dalfopristin (43.3%) and erythromycin (19.9%) were also observed. The overall multidrug resistant (MDR) rate was 14.9%. However, all GAS isolates were 100% sensitive to linezolid. M phenotype was observed in 85% of erythromycin resistant GAS isolates and mef (A) gene was identified in 96.9% of the erythromycin non susceptible GAS isolates. Among tetracycline resistant genes tet(M) was identified in 98.2% of the tetracycline non susceptible GAS isolates. Conclusion: In this study, we have identified 121 (60.2%) of the GAS isolates had at least one or more types of virulence factors and again more than half of the isolates,156 (77.6%) were resistant to one or more types of antibiotic classes. We also found that, an emerging penicillin non susceptible GAS isolates contrary to the previous reports and this might be an alarming condition. The most common erythromycin resistance mechanism in our study was efflux pumping which was encoded by the mef(A) gene where as that of tetracycline resistance mechanism was target site modification by ribosomal protection protein which was encoded by the tet(M) gene. Linezolid may be one of the promising drugs for the treatment of drug resistant GAS strains. The identification of different virulence factors and the increasing trends of drug non susceptibility especially to penicillin among GAS isolates requires a concern for continuous surveillance of the pathogen. Key Words: Streptococcus pyogenes, Group A Streptococci, antibiotic sensitivity testing, drug resistance genes, virulence factor, toxin, superantigen, Protease, DNase, EthiopiaItem Performance Evaluation of the First-Generation (1G) Color Plate Drug Susceptibility Testing Assay Identifying Drug- Resistant Mycobacterium Tuberculosis Isolates for Selected Anti- Tuberculosis Drugs(Addis Ababa University, 2024-03-13) Binyam Mebrat; Alem miritTuberculosis is a major global public health concern and the emergence of drug- resistant (DR)-TB has been a major challenge to the control of the disease, especially in developing countries, including Ethiopia. Despite availability of appropriate diagnostic tools for Mycobacterium tuberculosis drug susceptibility testing, accessibility and timely diagnosis of DR-TB in resource-constrained settings, such as Ethiopia, is often a challenge. The first- generation color plate assay (1G) is ideally suited for low resource settings due to low cost, minimal technical expertise needed, and rapid provision of DST results. Conclusion: The phenotypic DST 1G test is considered inexpensive (~$1.75), easy to use, rapid for identifying DR-TB using archived pulmonary and extra pulmonary MTB isolates from sputum and FNA and thus, it may be a viable option for DST in high endemic and resource limited settings like Ethiopia. The 1G test concurrently performs TB growth detection and DST in stored isolate’s, enabling timely treatment and preventing spread of DR-TB within the community. Additionally, the 1G test demonstrated good performance in detecting FNA (extra pulmonary) isolates, suggesting that it could be a useful alternative method for extra-pulmonary TB and DST diagnosis. Key Words: 1G test, color plate test, indirect proportional DST Method, Multidrug-resistant tuberculosis, Pre-XDR Tuberculosis, TuberculosisItem Prevalence and Antimicrobial Resistance Profile of Methicillin-Resistant Staphylococcus aureus (MRSA) and Factors Associated with it Among Patients with Wound Infections at Werabe Comprehensive Specialized Hospital, Central Ethiopia Region, Ethiopia(Addis Ababa University, 2024-05-12) Nejato Ousman; Alem AbrhaBackground: Staphylococcus aureus is a highly significant pathogen for global public health because of its extensive distribution, potential to cause infections that could be fatal, and continuous development of drug resistance. In settings such as the community and health care institutions, it is among the frequent causes of wound infection. Worldwide, the occurrence of drug-resistant S. aureus, particularly MRSA, has surged at a concerning rate. Nonetheless, prior research in Ethiopia and other nations found notable regional differences in the incidence and associated variables. Objective: To determine MRSA's prevalence and pattern of antimicrobial susceptibility as well as to assess factors associated with MRSA among patients with wound infection at Werabe Comprehensive Specialized Hospital, Werabe City, CER, Ethiopia. Methods: A cross-sectional study centered in a hospital was carried out from June 2023 to February 2024 G.C. S. aureus was isolated from a total of 384 wound swabs. Standard laboratory techniques were used to identify and culture S. aureus. For the identification of MRSA, testing for antimicrobial susceptibility was carried out in accordance with CLSI guidelines. Socio demographic information as well as other potential contributing factors was gathered using a structured questionnaire. SPSS version 27 was used for analysis once the data were entered into Epi-info. The study employed descriptive statistics and the Logistic regression analysis, both multivariate and bivariate. To evaluate the association between the variables that are dependent and independent, the odds ratio and 95% confidence interval were calculated, with a P value of less than 0.05 deemed statistically significant. Results: Of the 384 participants in the study with wound infections, 90 (23.4%, 95% CI: 19.3-28.0) had S. aureus isolated from them. Twenty-four (26.7%, 95% CI: 19-37) of the 90 S. aureus isolates were found to be MRSA. While the majority of the isolates (87.8%) were resistant to penicillin, Linezolid worked against every strain of S. aureus. All 24 of the isolates of MRSA had penicillin resistance. All (100%) of the MRSA isolates and twenty-eight (31.1%) isolates of S. aureus were multidrug-resistant (MDR). Various characteristics were evaluated for potential association with MRSA; however, only hospital stays longer than 72 hours (P=0.031, AOR: 3.06, 95% CI: 1.11 8.47) showed statistically significant association with MRSA Conclusion: The study showed significant prevalence of S. aureus (23.4%) and MRSA (26.7%) in patients with wound infection at WCSH. Both S. aureus and MRSA showed the highest resistance to penicillin but 100% sensitivity to linezolid. Key words: Wound infection, S. aureus, MRSA, MDR, WerabItem Molecular Epidemiology and Genetic Diversity of Epstein - Barr virus among lymphoma patients in Ethiopia(Addis Ababa University, 2024-06-13) Seifegebriel Teshome; Tamrat Abebe;Background: The Epstein-Barr virus (EBV) is a member of the herpesvirus family and is highly prevalent among the global population. It is estimated that more than 90% of adults have been exposed to this virus, making it one of the most common human viruses worldwide. After initial infection, infectious mononucleosis (IM), which typically occurs during childhood or adolescence, EBV establishes a long-lasting latent infection within B lymphocytes. EBV is associated with various diseases such as Burkitt's lymphoma, Hodgkin's lymphoma, nasopharyngeal carcinoma, and certain types of Gastric cancer. Studying the overall viral molecular epidemiology and genetic diversity of EBV can provide valuable insights into the pathogenesis, diagnosis, and management of EBV-associated diseases. The objectives of the study were on various aspects of EBV among lymphoma patients. It encompasses the detection of EBV, determining the genotypes, analysis of its methylation status, and identification of sequence variations. Methods: The study was conducted at Tikur Anbessa Specialized Hospital in Addis Ababa, Ethiopia, spanning from June 2021 to August 2023. A total of 305 participants were enrolled retrospectively and prospectively, and formalin-fixed paraffin-embedded (FFPE) tissues, blood, and lymph node biopsy samples were collected. From the whole blood samples, serum plasma and polymorphonuclear blood mononuclear cells (PBMCs) were isolated. Additionally, lymphomononuclear cells (LMNCs) were isolated from the lymph node biopsy samples. DNA was extracted from FFPE samples, PBMCs, and LMNCs. Serological tests were conducted using Viral capsid antigen (VCA) IgG to detect antibodies, and molecular detection and quantification were carried out using quantitative PCR (qPCR) by targeting EBV EBNA1 gene. Genotyping was performed using conventional PCR targeting the EBNA3C gene. The viral methylation profile was assessed using the IPLEX assay, and sequencing of the viral genome was performed using next generation sequencing technology (Illumina). We utilized statistical analysis tools such as descriptive statistics (mean, median, frequencies, percentages) and employed techniques like chi square test, bioinformatics, and phylogenetic tree analysis for in-depth data examination. SD, 17.5). Results: The mean age of the study participants was 38 years ( Among the lymphoma patients in the study, 32% (n=91) were diagnosed with Hodgkin lymphoma, while the remaining (68%) patients had non-Hodgkin lymphoma (n=197). Within the non-Hodgkin lymphoma cases, the highest percentage (18%) was attributed to small lymphocytic lymphoma, followed by diffuse Conclusion: This study presents the first evidence-based data in Ethiopia, providing valuable insights into the epidemiology and virological characteristics of lymphoma patients with EBV infection. These findings significantly contribute to our understanding of the relationship between EBV and lymphoma, emphasizing the importance of viral load, genotyping, methylation status, and genetic variations in different EBV genotypes. However, further research across diverse populations is necessary to broaden our knowledge of this complex virus and its association with lymphoma development. Such knowledge holds great potential for the development of region specific diagnostic and therapeutic strategies targeting EBV-associated lymphoma.