Hotspots of Multidrug-Resistant Bacteria: Sewage, Soil, and Cow Dung as Underestimated Drivers of Community AMR in Saurashtra region of Western India

Antimicrobial resistance (AMR) has emerged as a major public health threat, with environmental compartments increasingly recognized as important reservoirs and dissemination routes for resistant bacteria and resistance genes. This study investigated the occurrence, enumeration, and antibiotic resistance profiles of bacteria isolated from three highly contaminated environmental matrices in Rajkot, Gujarat, India: sewage water, sewage-contaminated soil, and fresh cow dung. Samples were processed in triplicate using standard spread-plate technique on Nutrient Agar. A total of 78 pure isolates were obtained and subjected to Gram staining, biochemical tests (IMViC, catalase, starch hydrolysis, nitrate reduction, urease, and gelatin hydrolysis), and antibiotic susceptibility testing against 18 clinically important antibiotics using the Kirby-Bauer disk diffusion method (CLSI M100, 2023 guidelines). Experiments were performed in biological triplicates and results expressed as mean ± SD. Multidrug-resistant (MDR) isolates were frequently encountered, particularly among Enterobacterales and Staphylococcus spp. The highest bacterial load was recorded in cow dung (7.8 ± 0.9 × 106 CFU/g), followed by sewage soil and sewage water. Resistance to third-generation cephalosporins, fluoroquinolones, and aminoglycosides was widespread. Identified genera included Escherichia coli, Enterobacter spp., Staphylococcus aureus, and Bacillus spp. This work provides the first comprehensive dataset on environmental AMR in Rajkot region and highlights the role of untreated sewage and animal manure as critical hotspots driving community-level AMR dissemination in India.