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GeneXpert is an effective and rapid molecular system used for tuberculosis (TB) diagnosis. It is expected to improve the detection rate and treatment outcomes needed to meet the sustainable development goals (SDG) and End TB strategy targets set for 2030. This study aimed to evaluate the impact of GeneXpert on diagnosis and anti-TB treatment outcomes in the post-millennium development goals (MDGs) in the capital city of Ethiopia. Hence, the global priority indicator based on the End TB Strategy for TB treatment success rate was met early in 2018 in Addis Ababa, Ethiopia, which was anticipated to be met by 2025. A retrospective health facilities-based study was conducted in Addis Ababa, Ethiopia. Records of all TB cases diagnosed and treated in selected health facilities from January 1.sup.st, 2015 to December 31.sup.st, 2018 were reviewed and included in the study. Data analysis of descriptive and inferential statistics was conducted using SPSS version 20. The reviewed records have shown that a total of 45,158 presumptive pulmonary TB (PTB) cases had accessed TB diagnosis services. Of which, 28.9% (13072/45158) were tested by AFB microscopy and 71.1% (32086/45158) were tested by GeneXpert. During the study period, the coverage of Xpert MTB/RIF testing increased to 94.9% in 2018 compared to 1.6% in 2015. The number of presumptive PTB cases tested with the GeneXpert system showed a significant increase compared to smear microscopy. The odds of positivity were detected in males compared to females. The odds of detecting TB cases were much higher among study participants aged 15-44 years compared to younger than 15 years. Treatment success rate showed a relative improvement each year between 2015 and 2018 with a mean of 92.6%. Reduced odds of treatment successes were observed in age categories older than 35 years, and in TB/HIV co-infected patients. Increased odds of treatment successes were reported in the years between 2016 and 2018 compared to 2015. Scaling up the Xpert MTB/RIF assay as a point-of-care test for presumptive TB cases in resource-limited settings would have a significant impact to meet the SDG and End TB strategy both in TB detection and treatment success rates.

Background Molecular surveillance of malaria and drug resistance monitoring typically uses dried blood samples (DBS) on filter papers. However, the use of Rapid Diagnostic Test (RDT) kits presents a promising yet unexplored alternative DNA source. This study aimed to assess the efficiency of DNA recovery from used RDT kits for molecular malaria surveillance and drug resistance monitoring by comparing its performance with the conventional filter paper DBS method. Methods Four hundred seventeen paired samples of RDT kits and DBS on filter paper were collected from malaria-positive cases at six health centres in the Gamo Zone of southern Ethiopia. DNA was extracted from both sample types using the Chelex-100 method, followed by nested polymerase chain reaction (PCR) targeting the 18S rRNA gene. Amplification of the 65 paired sub-samples of RDT and DBS-extracted DNA was carried out for the anti-malarial drug resistance gene, pfmdr1 . Nested PCR results from the two sample sources were compared using a 2 by 2 contingency table, along with Positive Percent Agreement (PPA) and Cohen’s Kappa agreement analysis. Results Of 417 paired samples, 391 (93.8%) of the DBS-extracted samples and 349 (83.7%) of the RDT-extracted samples were positive for malaria parasites using nested PCR. The PPA of samples extracted from RDT for P. falciparum detection was 89.2% (95% CI 84.7–92.5), while it was 81.9% (95% CI 74.6–87.6) for P. vivax. The overall Kappa value for agreement between nested PCR results from DBS and RDT-extracted samples was 0.64 (P < 0.001).This agreement was more pronounced (Kappa = 0.75; P < 0.001) in patients with high parasite load (> 100,000/μL). The amplification success rate for the pfmdr1 gene was 96.7% (63/65) for RDT kit samples and 100% (65/65) for DBS samples. Conclusion Used RDT kits can serve as an alternative DNA source and may be utilized for molecular surveillance of malaria and monitoring drug resistance in conditions when DBS is not available.

Detecting Plasmodium parasite DNA and host blood meals in the abdomens of fed mosquitoes enables timely decisions for effective malaria control. Hence, this study aimed to detect the Plasmodium parasite DNA and identify sources of blood meals in the abdomens of freshly fed Anopheles mosquitoes in South Ethiopia. A cross-sectional entomological survey was conducted in 12 malaria-endemic Kebeles near the southern Rift Valley Lakes, Abaya and Chamo, as a baseline for a trial evaluating house screening and/or ivermectin treatment for domestic animals to control malaria. Four hundred sixteen houses were selected for the Centers for Disease Control and Prevention (CDC) light trap collections. The mosquito species were identified using morphological and molecular methods. The polymerase chain reaction (PCR) technique was employed for the detection of Plasmodium parasite DNA and blood source identification. Plasmodium vivax (2.0%; 9/446) and P. falciparum (0.2%; 1/446) were detected in the abdomens of mosquitoes. Out of 446 tested Anopheles specimens, 85% (378/446) fed on six vertebrate hosts, while 15% (68/446) tested negative for these hosts. Goat blood was the most common, accounting for 67% (299/446), followed by human blood (23%; 102/446). Cow blood was identified in 19% (84/446) of the fed mosquitoes, while dog blood was present in 13% (57/446). Chicken and pig blood accounted for < 1% of blood meal sources. The overall human blood meal index was 23%, including mixed blood meal sources. Anopheles arabiensis was the most prevalent species, comprising 85% (380/446) of the nine Anopheles species identified, followed by An. pharoensis 7% (33/446) and An. tenebrosus 3% (13/446). Detecting Plasmodium parasites in the mosquitoes’ abdomens can enhance preparedness for malaria control. Anopheles mosquitoes obtain blood from various sources, with goats being the most common, followed by humans. Interventions targeting these mosquitoes may prioritise animal sources.

The emergence and rapid spread of coronavirus disease 2019 (COVID-19), a potentially lethal disease caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), is causing public health issues around the world. In resource-constrained nations, rapid Abbott SARS-CoV-2 antigen test kits are critical for addressing diagnostic gaps in health institutions and community screening. However, there is no evidence or proof of diagnostic performance in Ethiopia. The aim of this study was to compare the performance of PanbioTM Abbott SARS-CoV-2antigen rapid test kit to the gold standard, RT-PCR, in COVID-19 patients with clinical symptoms suggestive of COVID-19. A prospective, cross-sectional study was conducted between November 2021 and April 2022, on 120 suspected patients recruited from outpatient, emergency, and intensive care units in one of the tertiary hospitals in Ethiopia. Nasopharyngeal swabs were collected from suspected cases and were tested using the Abbott SARS-CoV-2 kit, a rapid diagnostic test (RDT) and compared to the reference standard RT-PCR. The sensitivity and specificity of the RDT were 74.2% and 100%, respectively. A total of 62 samples (51.6%) were RT-PCR positive. Of these, 46 were Ag-RDT positive. Sensitivity among symptomatic patients was 79.4% (95% CI 68.3-90). The Abbot RDT and RT-PCR had a Kappa value of agreement of 0.735 (p < 0.001). These values were acceptable when compared to the WHO's suggested thresholds. The finding from this study support the use of the Abbot RDT as a diagnostic tool in COVID-19 suspects, mainly in those with higher viral loads.

Background Efforts to control malaria face several challenges, including the presence of parasite reservoirs within the community that contribute to disease transmission. This study aimed to assess malaria prevalence among community members using microscopy and nested polymerase chain reaction (PCR) to detect parasite reservoirs in malaria-endemic settings. Methods A community-based cross-sectional study was conducted in malaria-endemic districts of the Gamo Zone in the southern Rift Valley of Ethiopia. The research was conducted as part of a baseline assessment for a trial evaluating the impact of house screening and ivermectin treatment of domestic animals on malaria incidence. Capillary blood samples were collected from the study participants to prepare blood films and dried blood spots. Malaria parasite detection and species identification were performed using both microscopy and nested PCR. Results Of 4745 participants screened by microscopy, malaria prevalence was 2.7% (126/4745; 95% CI: 2.2–3.2%). Nested PCR was performed on 1183 samples, including 97 microscopy-positive and 1,086 microscopy-negative samples. PCR confirmed 67% (65/97) of microscopy-positive cases, yielding a PCR-corrected microscopy prevalence of 1.4% (65/4745). Submicroscopic infections were detected in 6.1% (66/1086) of microscopy-negative samples. Malaria prevalence across Kebeles ranged from 0.5–2.3% by microscopy and 4.1–9.6% by PCR. Species misclassification by microscopy was common: 19.5% of P. falciparum infections were misidentified as P. vivax , 14.0% of P. vivax as P. falciparum , and 33.0% of microscopy-positive samples were PCR-negative. Conclusion These findings emphasize that community-based malaria diagnosis using microscopy underestimates malaria prevalence, highlighting the need for improved diagnostic methods.

Background Artemether–lumefantrine (AL) has been the primary anti-malarial drug used to treat uncomplicated Plasmodium falciparum malaria in Ethiopia since 2004. However, there have been recent reports of AL resistance mutations in different African countries, including Ethiopia. This is concerning and requires periodic monitoring of anti-malarial drug resistance. Therefore, the current study aimed to evaluate the therapeutic efficacy of AL in treating uncomplicated P. falciparum malaria in the Arba Minch Zuria District, Gamo Zone, Southwest Ethiopia. Methods A single-arm prospective study with a 28-day follow-up period was conducted from July to October 2022. Capillary blood samples were collected for RDT and microscopic examination. The study enrolled monoinfected P. falciparum patients aged ≥ 18 years at Ganta Sira Health Post. Sociodemographic and clinical data were recorded, and a dried blood spot (DBS) was prepared for each participant. Nested polymerase chain reaction (nPCR) genotyping of the msp-1 and msp-2 genes was only performed for recurrent cases to distinguish between recurrence and reinfection. Data entry and analysis were performed using the WHO Excel spreadsheet and SPSS version 26. Results A total of 89 patients were enrolled, and 67 adequately completed the 28-day follow-up period. AL showed a 100% clearance rate for fever on day 2 and asexual parasites on day 3. Gametocytes were detected in 13.5% (12/89) of the participants. The gametocyte clearance rate was 58.3% (7/12) until day 7 and 100% (12/12) until day 14. Five participants developed recurrent malaria, three of whom experienced relapse and two of whom experienced reinfection. Based on the Kaplan–Meier survival analysis, the PCR-uncorrected and PCR-corrected cumulative incidence of success were 93.7% (95% CI 85.5–97.3) and 96.2% (95% CI 85.5–98.7), respectively. Conclusion AL was efficacious in treating uncomplicated P. falciparum malaria in the study area. However, the detection of recurrent patients highlights the need for continuous efficacy studies in this area.

The persistence of tuberculosis (TB) infection in some patients after treatment has highlighted the importance of drug susceptibility testing (DST). This study aimed to determine the drug susceptibility patterns of Mycobacterium tuberculosis (M. tuberculosis) isolates from pulmonary TB (PTB) patients in Central and Southern Ethiopia. A health institution-based cross-sectional study was conducted between July 2021 and April 2022. Sputum samples were collected from newly diagnosed smear microscopy and/or Xpert MTB/RIF-positive PTB patients. The samples were processed and cultivated in Lowenstein-Jensen (LJ) pyruvate and glycerol medium. M. tuberculosis isolates were identified using polymerase chain reaction (PCR) based region of difference 9 (RD9) deletion typing. Phenotypic DST patterns of the isolates were characterized using the BACTEC MGIT™ 960 instrument with SIRE kit. Isoniazid (INH) and Rifampicin (RIF) resistant M. tuberculosis isolates were identified using the GenoType® MTBDRplus assay. Sputum samples were collected from 350 PTB patients, 315 (90%) of which were culture-positive, and phenotypic and genotypic DST were determined for 266 and 261 isolates, respectively. Due to invalid results and missing data, 6% (16/266) of the isolates were excluded, while 94% (250/266) were included in the paired analysis. According to the findings, 14.4% (36/250) of the isolates tested positive for resistance to at least one anti-TB drug. Gene mutations were observed only in the rpoB and katG gene loci, indicating RIF and high-level INH resistance. The GenoType® MTBDRplus assay has a sensitivity of 42% and a specificity of 100% in detecting INH-resistant M. tuberculosis isolates, with a kappa value of 0.56 (95%CI: 0.36-0.76) compared to the BACTEC MGIT™ DST. The overall discordance between the two methods was 5.6% (14/250) for INH alone and 0% for RIF resistance and MDR-TB (resistance to both INH and RIF) detection. This study reveals a higher prevalence of phenotypic and genotypic discordant INH-resistant M. tuberculosis isolates in the study area. The use of whole-genome sequencing (WGS) is essential for gaining a comprehensive understanding of these discrepancies within INH-resistant M. tuberculosis strains.

Background Identification of non-sputum diagnostic markers for tuberculosis (TB) is urgently needed. This exploratory study aimed to discover potential serum protein biomarkers for the diagnosis of active pulmonary TB (PTB). Method We employed Proximity Extension Assay (PEA) to measure levels of 92 protein biomarkers related to inflammation in serum samples from three patient groups: 30 patients with active PTB, 29 patients with other respiratory diseases with latent TB (ORD with LTBI+), and 29 patients with other respiratory diseases without latent TB (ORD with LTBI-). To understand the functional mechanisms associated with differentially expressed proteins, we performed Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses. Least absolute shrinkage and selection operator (LASSO) regression was employed to identify potential TB diagnostic protein biomarkers. Network interactions among the identified candidate diagnostic markers were then analyzed, and their diagnostic performance was evaluated using logistic regression and receiver operating characteristic (ROC) analysis. Result The analysis revealed 37 differentially expressed proteins (DEPs) in the active PTB group compared to both ORD with LTBI + and ORD with LTBI- groups. Gene Ontology analysis indicated that these DEPs were primarily involved in the inflammatory response, while KEGG enrichment analysis highlighted the cytokine-cytokine receptor interaction pathway as the top significant hit. LASSO regression identified eight promising candidate protein biomarkers: IFN-gamma, LIF, uPA, CSF-1, SCF, SIRT2, 4E-BP1, and GDNF. The combined set of these eight proteins yielded an AUC of 0.943 for differentiating active PTB from ORD with LTBI+, and an AUC of 0.927 for distinguishing PTB from ORD with LTBI-. Conclusion We have identified eight protein markers that reliably differentiate active PTB from ORD irrespective of LTBI presence. Further large-scale validation and translation of these protein markers into a user-friendly and affordable point-of-care test hold the potential to significantly enhance TB control in high-burden regions.

Background Tuberculosis (TB) is caused by Mycobacterium tuberculosis complex (MTBC). Mapping the genetic diversity of MTBC in high TB burden country like Ethiopia is important to understand principles of the disease transmission and to strengthen the regional TB control program. The aim of this study was to investigate the genetic diversity of Mycobacterium tuberculosis complex (MTBC) isolates circulating in the South Omo, southern Ethiopia. Methods MTBC isolates ( N  = 156) were genetically analyzed using spacer oligotyping (spoligotyping) and mycobacterial interspersed repetitive unit-variable number of tandem repeat (MIRU-VNTR) typing. Major lineages and lineages were identified using MTBC databases. Logistic regression was used to correlate patient characteristics with strain clustering. Results The study identified Euro-American (EA), East-African-Indian (EAI), Indo-Oceanic (IO), Lineage_7/Aethiops vertus, Mycobacterium bovis and Mycobacterium africanum major lineages in proportions of 67.3% (105/156), 22.4% (35/156), 6.4% (10/156), 1.9% (3/156), 1.3% (2/156) and 0.6% (1/156), respectively. Lineages identified were Delhi/CAS 23.9% (37/155), Ethiopia_2 20.6% (32/155), Haarlem 14.2% (22/155), URAL 14.2%(22/155), Ethiopia_3 8.4% (13/155), TUR 6.5% (10/155), Lineage_7/Aethiops vertus 1.9% (3/155), Bovis 1.3% (2/155), LAM 1.3% (2/155), EAI 0.6% (1/155), X 0.6% (1/155) and Ethiopia H 37 Rv-like strain 0.6% (1/155). Of the genotyped isolates 5.8% (9/155) remained unassigned. The recent transmission index (RTI) was 3.9%. Orphan strains compared to shared types (AOR: 0.09, 95% CI: 0.04–0.25) were associated with reduced odds of clustering. The dominant TB lineage in pastoral areas was EAI and in non-pastoral areas was EA. Conclusion The epidemiological data, highly diverse MTBC strains and a low RTI in South Omo, provide information contributing to the TB Control Program of the country.

Background Plasmodium falciparum and Plasmodium vivax are coendemic in Ethiopia, with different proportion in different settings. Microscopy is the diagnostic tool in Ethiopian health centres. Accurate species-specific diagnosis is vital for appropriate treatment of cases to interrupt its transmission. Therefore, this study assessed the status of species-specific misdiagnosis by microscope compared with polymerase chain reaction (PCR). Methods A health facility based cross-sectional study was conducted from November 2019 to January 2020 in Kolla Shelle Health centre, Arba Minch Zuria district. The study population were suspected malaria cases, who visited the health centre for a diagnosis and treatment. Consecutive microscopy positive cases as well as a sample of microscopically negative cases were included for molecular analysis by polymerase chain reaction (PCR). Results 254 microscopically negative and 193 microscopically positive malaria suspects were included. Of the 193 malaria positive cases, 46.1% [95% confidence interval (CI) 38.9–53.4] (89/193) were P. falciparum infection, 52.3% (95% CI 45.0–59.5) (101/193) were P. vivax infection, and 1.6% (3/193) had mixed infection of P. falciparum and P. vivax . Of the microscopically positive cases of P. falciparum , 3.4% (3/89) were P. vivax and 11.2% (10/89) were mixed infections with P. falciparum and P. vivax and a single case was negative molecularly. Similarly, of the microscopically positive P. vivax cases, 5.9% (6/101) were P. falciparum and 1% (1/101) was mixed infection. Single case was negative by molecular technique. Of the 254 microscopically negative cases, 0.8% were tested positive for P. falciparum and 2% for P. vivax by PCR . Considering molecular technique as a reference, the sensitivity of microscopy for detecting P. falciparum was 89.2% and for P. vivax , it was 91.2%. The specificity of microscopy for detecting P. falciparum was 96.1% and for P. vivax , it was 97.7%. However, the sensitivity of microscopy in detecting mixed infection of P. falciparum and P. vivax was low (8.3%). Conclusion There were cases left untreated or inappropriately treated due to the species misidentification. Therefore, to minimize this problem, the gaps in the microscopic-based malaria diagnosis should be identified. It is recommended to regularly monitor the competency of malaria microscopists in the study area to improve species identification and diagnosis accuracy.