Background: Persistence and dissemination of pathogenic bacteria and antibiotic resistancerngenes (ARGs) in aquatic environments remain a serious public health issue. The aquaticrnenvironment may serve as a key transmission pathway for pathogenic bacteria and ARGs tornand from humans. Continual faecal discharge into the environments, together with high levelsrnof chemical and pharmaceutical pollutants are creating conditions favourable for thernpersistence and spread of antibiotic resistant pathogens. The aim of the study was torninvestigate the distribution of ARGs, and functional and molecular characteristics ofrnpotentially pathogenic bacteria in the Akaki river that flows through Addis Ababa, Ethiopia,rnand its toxicogenomic effect in model organisms. rnMethods: Water and sediment samples were collected from five sites along the Akaki riverrnsystem for isolation of potential bacterial pathogens and detection of ARGs. The diversity andrnabundance of 84 ARGs and 116 clinically important bacteria were evaluated using DNArnqPCR analysis. Potential bacterial pathogens were isolated using selective agar techniquesrnand identified by MALDI-TOF. Isolates were analyzed for their resistance to different classesrnof antibiotics and further characterized by PCR and whole-genome sequencing. Therntoxicogenomic effect of the Akaki river water was also assessed by gene expression analysisrnof selected genes in exposed Caenorhabditis elegans and Pseudomonas aeruginosa PAO1. rnResults: Genes associated with fluoroquinolone, aminoglycoside, Î²-lactamase, macrolide,rnmultidrug resistance efflux pumps, tetracycline efflux pumps, and vancomycin resistancernwere detected in the water and sediment samples. The majority of ARGs were identified fromrnsites in close proximity to anthropogenic activities such as hospitals, irrigation systems, andrnindustries. Aminoglycoside acetyltransferase (AAC(6)-Ib-cr), aminoglycoside adenylyltransferasern(aadA1),rnÎ²-lactamasern(blaOXA-10rn),rn quinolone resistance S (qnrS), macrolide effluxrnprotein A (mefA) and tetracycline resistance (tetA) genes were detected at all sampling sites.rnSurprisingly, much fewer ARGs were detected in the associated sediments, and the sedimentrncollected from the hospital site had the highest diversity and level of resistance genes. Genesrnsuch as blaOXA-10rn, blaOXA-2rn, aadA, ermC and oprM were persistent in all sediment samples.rnThere was significant variation in the abundance of ARGs between study sites, with those near the hospitals having the highest abundance in both waters and sediments. Escherichiarncoli strains isolated from the Akaki river were most frequently resistant to erythromycinrn(97%), doxycycline (78.5%), tetracycline (75.6%), and amoxicillin (75.6%). The overallrnprevalence of resistance to cephalosporin and carbapenem was 7.9% and 5%, respectively andrn80.9% of E. coli isolates were MDR. Most E. coli isolates (70.8%) harbored enteroaggregativernheat-stable toxin 1 (EAST1) gene, whereas the heat-labile toxin (LT) was detected in only 34rn(20.2%) of the isolates. The prevalence of Shiga toxin-producing E. coli was 7.7%. The E.rncoli isolates belonged to 20 different sequence types, with ST10, ST69 and ST361 beingrnpredominant. The Î²-lactamase genes were the commonly detected resistance genes in E. coli.rnThe most prominent plasmid sequence replicons detected were from the IncF family (65%),rnCol (10%), and IncX3 (7.5%). The Aeromonas spp. isolated from the Akaki river werernresistant to a wide range of antibiotics with the highest being resistant to amoxicillin (144,rn100%), ampicillin (142, 99%), amoxicillin/clavulanate (117, 81%), imipenem (75, 52%),rnertapenem (132, 92%) and doxycycline (134, 93%). The heat-labile cytotonic enterotoxin (alt)rnwas the predominant toxin gene detected, followed by a heat-stable cytotonic enterotoxinrn(ast). Almost all Aeromonas isolates in the current study were new sequence types. P.rnaeruginosa isolates detected in the Akaki river were resistant to different classes of antibioticsrnand resistance is mediated by diverse group of ARGs with the majority were multi-drug effluxrnsystems. Akaki river water induced a significant change in gene expression in exposed C.rnelegans and P. aeruginosa PAO1. Although the levels of almost all analyzed metals werernbelow the permissible limit, the compiled effect resulted in transcriptional changes at thernmolecular level. A significant spatial variation was observed between the sampling points inrnthe expression of certain genes. rnConclusion: The findings suggest that the water phase, rather than the sediments in the Akakirnriver, are a potential conduit for the spread of ARGs and thus antibiotic resistant bacteria. Thernoccurrence of antibiotic resistance in E. coli, Aeromonas spp. and P. aeruginosa isolated fromrnAkaki river provides evidence for the need to develop prevention and control strategies tornlimit the spread of antibiotic-resistant bacteria in the aquatic environment. The presence ofrnvirulent E. coli and Aeromonas spp. in the water could be posing a serious health risk to the public. The Akaki river water has selection pressure in bacteria and elicits a toxic effect in C.rnelegans at the molecular level result in a higher risk of infectious disease.