Wastewater Metagenomic Reanalysis of Antibiotic Resistance Genes in Public Datasets from Türkiye (Ankara and Hatay)

Background/Objectives: Antimicrobial resistance in microbial communities is a global health concern that leads to millions of deaths each year. Many bacterial pathogens have resistance to multiple antibiotics. Domestic wastewater treatment facilities are reservoirs for antibiotic-resistant bacteria and resistance genes. Wastewater-based epidemiology surveillance is crucial for monitoring antibiotic resistance genes (ARGs). Türkiye has one of the highest levels of antibiotic resistance with a lack of research on resistomes. This study is a focused reanalysis of publicly available wastewater metagenomes from Türkiye, comparing them to global and other country's results. Methods: Ten metagenomic data of wastewater treatment from Türkiye were downloaded from NCBI-SRA database. Metagenome assemblies were performed and high-quality metagenome-assembled genomes (HQ-MAGs) were included in the study. Taxonomic annotations and antibiotic resistance profiles were identified in both the metagenome assemblies and HQ-MAGs. Results: A total of 401 different ARGs in 25 antibiotic classes have been identified, including Mcr (including mcr-1, mcr-2, mcr-3 and mcr-5 variants) and optrA. The vanR two-component regulatory system genes for controlling vancomycin antibiotic resistance were one of the most dominant along with other vancomycin resistance genes such as vanA and vanB. A total of 115 HQ-MAGs were obtained with at least eight ARGs. The HQ-MAG with the highest number of resistance genes (58) was found to belong to E. coli. The most frequently encountered resistance genes in HQ-MAGs were the multidrug ABC transporter, vanR, bacA and patA which confer resistance to multidrug, glycopeptide, bacitracin and fluoroquinolone antibiotic groups, respectively. Conclusions: To effectively address the problems of antibiotic resistance outbreaks, comparable AMR surveillance at national and global levels is required for the identification and prioritization of ARGs and resistance genes. This is the first report conducted in Türkiye.