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Background Newcastle disease virus (NDV) is a causative agent of Newcastle disease (ND), a major infectious poultry disease associated with significant economic losses. Vaccination is usually effective at preventing the disease. However, in Ethiopia, ND is commonly detected in both unvaccinated and vaccinated chickens. In this study, we aimed to evaluate the pathogenicity of NDV isolated from both vaccinated and unvaccinated chickens, as well as to compare the antigenicity of the isolates with vaccine strains and genotyping by using the F -gene sequence. Methods The partial F gene sequences of all isolates and the mean death times (MDTs) of representative isolates were used to determine genotype and pathogenicity of the isolates. Antigenicities were assayed with the hemagglutinin inhibition (HI) and virus neutralization (VN) tests using antiserum against the vaccine Hitchner B1 (HB1), which is the most commonly used NDV vaccine in Ethiopia. Thermostability was evaluated by incubating infected allantoic fluid at 56 °C. Results Out of 231 samples tested, 10.8% (25/231) were positive for virus isolation. The F gene cleavage sites of all 25 isolates had 112 RRQKRF 117 , a characteristic of virulent NDVs. The MDTs of representative isolates were less than 60 h, indicating highly virulent (velogenic) pathotypes. The HI test revealed significant differences between our isolates and the HB1 vaccine strain, but the VN test showed no antigenic difference. Phylogenetic analysis based on the partial F gene sequences showed that all the isolates belonged to sub-genotype VII.1.1 of genotype VII, which is closely related to NDV strains from the Middle East and Eritrea. Thermostability test showed two of the 25 isolates were thermostable. Discussion Although the HI test indicates antigenic differences between the velogenic Ethiopian isolates and the HB1 vaccine, the VN test showed that the vaccine could protect infections with these isolates. Phylogenetic analysis showed that all studied isolates belong to sub-genotype VII.1.1 of genotype VII, diverging from previously reported genotype XXI in Ethiopia. Conclusions In Ethiopia, NDV genotype VII 1.1 is widely distributed. Since these viruses showed the same antigenicity as the HB1 vaccine in VN test, the occurrence of ND in vaccinated chickens may be due to vaccine failure caused by inadequate management or immunosuppression due to other infectious diseases.

Background Arboviruses continue to threaten global health because of their rapid geographical expansion and significant disease burden. Of the over 500 recognized arboviruses, approximately 150 affect humans, and around 50 affect domestic animals and wildlife. The spread and impact of these viruses have increased significantly over the past three decades, driven by the proliferation of their vectors and the rise of global trade and travel. Methods In this study, we used molecular methods to characterize mosquito species diversity and host feeding preferences across Ethiopia’s Great Rift Valley. Mosquitoes were collected from diverse habitats in the Great Rift Valley of Ethiopia using Centers for Disease Control and Prevention (CDC) light traps, BG-Sentinel traps, and hand aspirators. The area was chosen for its high vector diversity, suitable breeding habitats, and the epidemiological importance of arboviruses. Morphological identification was conducted, and 204 blood-fed mosquitoes were selected. Genomic DNA was extracted, followed by polymerase chain reaction (PCR) amplification targeting the COI gene. Blood meal analysis was performed using vertebrate-specific primers targeting the 12S rRNA gene. Mosquito species identification, genetic diversity analysis, and phylogenetic analyses were conducted. Results Of 6601 collected mosquitoes, 4977 were identified morphologically, comprising 399 Aedes , 2861 Culex , 1841 Anopheles , and 275 Mansonia species. COI DNA barcode analysis identified 142 mosquito specimens belonging to 16 species, with Anopheles coustani , Cx. tritaeniorhynchus , Cx. pipiens complex, Mansonia africana , and Ma. uniformis being the predominant species. Blood meal analysis ( n  = 71 successful amplifications) revealed a primary reliance on humans and cattle. Cx. pipiens complex showed a strong anthropophilic tendency, while Cx. tritaeniorhynchus and Ma. uniformis exhibited broader host ranges. Genetic diversity indices showed significant Fu’s F s statistics for Cx. pipiens complex, Cx. tritaeniorhynchus , Ma. africana , and Ma. uniformis . Conclusions This study offers valuable preliminary insights into the diversity of mosquito species, genetic variation, and host-feeding preferences within the Ethiopian Rift Valley. The findings emphasize the potential of molecular techniques to enhance traditional entomological methods and improve the accuracy of mosquito identification. While the study is limited in both geographic and temporal scope, it highlights mosquito species of medical and veterinary significance and suggests implications for arboviral disease surveillance. Graphical Abstract

Bovine tuberculosis (bTB) is a threat to cattle health and public safety. The current control programs are hampered by wildlife reservoirs and socioeconomic barriers. Vaccinating cattle with Bacillus Calmette-Guérin (BCG) effectively reduces transmission, offering a potential solution for controlling bTB. A key requirement for vaccination strategies using BCG is the validation of defined antigens to differentiate infections among vaccinated animals (DIVA). We compared tuberculin with DIVA peptide cocktails (ESAT-6, CFP-10, and Rv3615c) in 67 unvaccinated and 67 BCG-vaccinated cattle exposed to M. bovis in a natural setting. The cattle were tested every 4 months with a skin test and every 2 months with interferon-gamma (IFN-γ) release assays (IGRA) over a year of exposure. Before exposure, the DIVA skin, DIVA IGRA, and tuberculin tests showed 100% specificity in unvaccinated control calves. After exposure, the DIVA skin, DIVA IGRA, and comparative cervical tuberculin (CCT) tests had comparable sensitivities of 46% (95% CI 36, 56), 45% (95% CI 35, 55), and 47 (95% CI 37, 57), respectively, when assessed against animals positive by M. bovis culture PCR. The results suggest that test-and-slaughter control strategies using tests with low sensitivity are not expected to be effective in controlling bTB in high-prevalence herds, and highlight an urgent need to improve the sensitivity of diagnostic tests for bTB in these settings.

Arboviral diseases transmitted by mosquitoes, including dengue, Chikungunya, Zika, yellow fever, West Nile virus (WNV), and Rift Valley fever (RVF), pose a significant public health challenge globally, particularly impacting populations in low and middle-income countries. Conventional mosquito control methods, which primarily rely on insecticides, face critical challenges, including the development of insecticide resistance and environmental concerns. In this context, Wolbachia , an endosymbiotic bacterium, presents an alternative strategy due to its ability to manipulate mosquito reproduction and impede the transmission of pathogens. This study aimed to detect and assess the genetic diversity of Wolbachia and prophage WO in Ethiopian mosquitoes. Mosquitoes were collected from various ecological niches in the Great Rift Valley. Molecular analyses were performed to identify the presence of Wolbachia using PCR targeting the 16 S rRNA and wsp genes. Additionally, the presence of prophage WO was assessed by detecting the conserved orf7 capsid protein gene. To understand genetic diversity, phylogenetic and genetic diversity analyses were performed. Wolbachia was detected in 44.2% (34/77) of mosquitoes using the 16 S rRNA gene and 46.8% (36/77) using the wsp gene. The highest prevalence was observed in Cx. pipiens complex (100%, 11/11) and Ma. uniformis (92.3%, 12/13). Prophage WO was detected in 46.8% (36/77) of mosquitoes, with evidence of multiple-strain co-infections in Cx. pipiens complex. Phylogenetic analysis classified all isolates within Wolbachia pipientis Supergroup B. This study provides the first preliminary characterization of Wolbachia and prophage WO in Ethiopian mosquitoes, revealing evidence of genetic diversity. These findings lay the conceptual foundation of potential Wolbachia -based vector control strategies in Ethiopia and underscore the need for further studies on strain-specific impacts on vector competence and arboviral transmission dynamics.

Background Peste des Petits Ruminants (PPR) is a highly contagious viral disease primarily affecting goats and sheep, with clinical manifestations ranging from peracute disease to subclinical infection, particularly in atypical hosts such as cattle. The role of atypical hosts such as cattle to the spread of PPR remains controversial, with conflicting reports in the literature. Despite its worldwide significance, considerable knowledge gaps exist regarding the pathogenesis and clinical progression in both primary and atypical hosts. This study aimed to elucidate the tissue tropism, pathogenesis, virus shedding, clinical progression, and pathology associated with experimental PPR virus infection in indigenous goats and cattle. To this end, 32 animals—16 goats and 16 cattle—were intranasally inoculated with the Ethiopia/Habru/2014 Lineage-IV strain of the PPR virus followed by detailed clinical evaluations and systematic sampling at pre-established intervals to assess serological conversion, viral shedding, and the pathogenesis of the infection across both species. Results The results show that goats exhibited typical clinical signs 4 days post-inoculation, with seroconversion by day 6 and early detection of viral RNA in swabs and tissues by day 3 and virus isolation starting day 4. In contrast, cattle exhibited minimal clinical signs, with seroconversion occurring at day 8 with viral RNA detected in tissue samples at day 4 and virus isolation starting day 6 in tissues and in a single nasal swab at day 8. Clinical scores and tissue positivity rates significantly differed between goats and cattle ( P  = 0.007 and P  < 0.001, respectively). While goats exhibited expected gross and histopathological lesions, cattle showed only nonspecific lesions. Conclusions Together, our findings highlight the importance of comparative pathology studies for better understanding virus dynamics and transmission pathways that may help inform more effective PPR control programs. Future research should explore the pathogenesis of different PPRV lineages in cattle, assessing variations in disease progression and potential for epidemiological impact.

Newcastle Disease (ND) is a highly contagious and fatal poultry disease caused by Newcastle disease virus (NDV) and has a global distribution. To control this disease, a rapid method for diagnosing it is needed. PCR-based molecular diagnostics such as real-time reverse transcription-PCR (RRT-PCR) for detecting NDV genes have been conducted worldwide. However, because PCR methods are time-consuming and require a well-equipped laboratory, they are not well-suited for use in field and resource limited areas. In this study, we established a field-friendly loop-mediated isothermal amplification (LAMP) assay for NDV diagnosis using primers targeting the highly conserved L gene, which bypasses the need for RNA extraction and makes it more suitable for field application than previously reported LAMP methods. Our LAMP method is capable of detecting a broad range of NDV genotypes and showed no cross-reactivity with other avian viral diseases or the host genome. The reaction is completed within 35 minutes of incubation at 65°C. NDV was successfully detected directly from swab and tissue samples without the need for conventional RNA extraction. The LAMP method developed in this study offers a rapid, affordable, and field-friendly diagnostic tool for NDV detection.

Background Pseudomonas species, including P. aeruginosa , P. putida , and P. fluorescens , are zoonotic bacterial pathogens responsible for significant disease and mortality in both farmed and wild fish worldwide. In Ethiopia, these bacteria have been identified in Sebeta fish ponds and Rift Valley lakes, yet there is limited data on their molecular and phenotypic characteristics in local aquaculture systems. To address this gap, a cross-sectional study was conducted from November 2022 to May 2023 in selected aquaculture ponds in Ethiopia. Results A total of 637 samples were aseptically collected from the muscle, liver, spleen, and kidney of fish using purposive sampling. Pseudomonas base agar selective medium morphological characteristic and biochemical tests were used to isolate and identify pseudomonas species. Pseudomonas species were isolated from 81 samples, representing 12.7% of the total. Among these isolates, 85.6% displayed virulence traits, including β- hemolysis on 5% sheep blood agar. Additionally, 75 isolates (92.59%) was confirmed using conventional PCR with Pseudomonas-specific primers. Of the PCR-positive samples, 8 (10.66%) were identified as P. aeruginosa , 28 (37.63%) as P. putida , and 39 (52%) as P. fluorescens from Nile Tilapia ( O. niloticus ). Antibiotic susceptibility testing on ten representative isolates revealed that all strains were sensitive to Ciprofloxacin, Gentamicin, and Ceftriaxone but resistant to Amoxicillin and Penicillin. Conclusions The findings indicate that Pseudomonas species carrying virulence genes, exhibiting β- hemolytic activity, and showing resistance to commonly used antibiotics in both human and veterinary medicine are present in aquaculture. The detection of this pathogen in 75 fish samples raises concerns about potential outbreaks and zoonotic transmission. Therefore, further research on the molecular epidemiology of the disease is necessary to understand inter-host transmission and antibiotic resistance patterns.

Wastewater surveillance systems have become an important component of COVID-19 outbreak monitoring in high-income settings. However, its use in most low-income settings has not been well-studied. This study assessed the feasibility and utility of wastewater surveillance system to monitor SARS-CoV-2 RNA in Addis Ababa, Ethiopia. The study was conducted at nine Membrane Bio-reactor (MBR) wastewater processing plants. The samples were collected in two separate time series. Wastewater samples and known leftover RT-PCR tested nasopharyngeal swabs were processed using two extraction protocols with different sample conditions. SARS-CoV-2 wastewater RT-PCR testing was conducted using RIDA GENE SARS-CoV-2 RUO protocol for wastewater SARS-CoV-2 RNA testing. Wastewater SARS-CoV-2 RNA RT-PCR protocol adaptation, optimization, and detection were conducted in an Addis Ababa, Ethiopia context. Samples collected during the first time series, when the national COVID-19 case load was low, were all negative. Conversely, samples collected during the second time series were all positive, coinciding with the highest daily reported new cases of COVID-19 in Ethiopia. The wastewater-based SARS-CoV-2 surveillance approach is feasible for Addis Ababa. The COVID-19 wastewater based epidemiological approach can potentially fill the evidence gap in distribution and dynamics of COVID-19 in Ethiopia and other low-income settings.

Introduction Peste des petits ruminants virus (PPRV) causes a highly devastating disease of sheep and goats, peste des petits ruminants (PPR), which is targeted for global control and eradication by 2030. The serological diagnostic tool kits for accurate diagnosis of PPR have inherent strengths and weaknesses that require parallel validation and optimization across animal species. Thus, the objective of this study was to evaluate diagnostic performance of haemagglutinin based PPR blocking ELISA (HPPR- b-ELISA), that was developed by Africa Union Pan African Veterinary Vaccine Center for specific detection of anti- PPRV antibodies. Methods In preliminarily investigation, diagnostic performance of the HPPR-b-ELISA®, commercial PPR competition ELISA (c-ELISA) and virus neutralization test (VNT) were compared for the detection of anti-PPRV antibodies in goats, sheep, cattle and camels. Results The sensitivity and specificity of HPPR- b-ELISA® were 79.55 and 99.74%, respectively, compared to c-ELISA. The HPPR- b-ELISA® was in perfect agreement (κ = 0.86) with the c-ELISA in all sera collected from goats, sheep and cattle. There was almost perfect agreement between the species of goats (κ = 0.82) and sheep (κ = 0.98), while the agreement was substantial in cattle (κ = 0.78) and no agreement was observed in camels (κ = 0.00). Similarly, the sensitivity and specificity of the HPPR b-ELISA were 80 and 96.36%, respectively compared to VNT with almost perfect agreement in goats (κ = 0.83) and sheep (κ = 0.89), moderate in cattle (κ = 0.50) and none in camels (κ = 0.00). Conclusion Our study revealed that HPPR- b-ELISA is a suitable and valid method that can alternatively be used for screening and monitoring of PPR in sheep, goats and cattle except for camels.

Lumpy skin disease (LSD) is a viral disease caused by LSD virus and is one of the most economically significant transboundary and emerging diseases of cattle. LSD causes considerable economic losses due to emaciation, damage to hides, infertility, and loss of milk production. In Ethiopia, the disease is distributed almost in all regions and is regarded as one of the most economically important livestock diseases in the country. An outbreak investigation of the disease was monitored from October 2016 to April 2017 in southern pastoral areas of Bale Zone, Oromia, Ethiopia. In December 2016, LSD outbreak occurred in Sawena district of Bale Zone, from which necessary biopsy samples were collected from actively infected animals for the purpose of virus isolation, and characterization using different molecular techniques at National Animal Health and Diagnostic Investigation Center (NAHDIC) of Sebeta, Ethiopia. In addition, clinical examination of infected and in-contact animals was carried out together with a questionnaire survey. Based on the clinical manifestations, LSD was recorded in 18% (94/522) of examined cattle, whereas biopsy samples from 20 clinically positive animals were collected for further laboratory process. The morbidity rate was higher in animals less than two years 28.97% (31/107) than other ages and showed a statistically significant difference with P<0.05. Female animals showed higher morbidity rate of 20.59% (76/369) than male animals (11.76%) (18/153) with a significant difference at P≤0.003. Mortality rate and case fatality were also significantly higher in young animals than other age groups. Viruses were isolated from both skin biopsies and nasal swabs on Vero cell line. From both skin biopsies and nasal swabs, the virus DNA was identified by amplifying the 172 bp DNA fragment using real-time and conventional PCR. Providing adequate diagnostic facilities, establishing strategic policies for effective control and eradication and awareness creations for communities for early identification or reporting were recommendations made to minimize economic losses of the disease.